i  

MOLECULAR DYNAMICS OF TUBER AND SEED YIELD IN 

Sphenostylis stenocarpa (Hochst. ex A. Rich. Harms) (AFRICAN YAM 

BEAN) 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

ALADELE, ANDREW KOLAWOLE 

(22PCO02401) 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

MARCH, 2025 



ii  

MOLECULAR DYNAMICS OF TUBER AND SEED YIELD IN 

Sphenostylis stenocarpa (Hochst. ex A. Rich. Harms) (AFRICAN YAM 

BEAN) 

 

 

 

BY 

 

 

 

 

ALADELE, ANDREW KOLAWOLE 

(22PCO02401) 

B.Agric Plant Physiology and Crop Production, Federal University of 

Agriculture Abeokuta 

 

 

 

 

 

 

A DISSERTATION SUBMITTED TO THE SCHOOL OF 

POSTGRADUATE STUDIES IN PARTIAL FULFILLMENT OF THE 

REQUIREMENTS FOR THE AWARD OF THE DEGREE OF MASTER 

OF SCIENCE (M.Sc.) IN BIOLOGY IN THE DEPARTMENT OF 

BIOLOGICAL SCIENCES, COLLEGE OF SCIENCE AND 

TECHNOLOGY, COVENANT UNIVERSITY, OTA, OGUN STATE 

NIGERIA 

 

 

 

 

 

 

 

 

 

 

MARCH, 2025 



iii  

ACCEPTANCE 

This is to attest that this dissertation has been accepted in partial fulfilment of the requirements 

for the award of the degree of Masters of Science in Biology, in the Department of Biological 

Sciences, College of Science and Technology, Covenant University, Ota, Ogun State. 

 

 

 

 

 

 

 

 

Miss Adefunke F. Oyinloye 

(Secretary, School of Postgraduate Studies)   Signature and Date 

 

 

 

 

 

 

 

 

 

 

Prof. Conrad A. Omonhinmin 

(Dean, School of Postgraduate Studies)   Signature and Date 



iv  

DECLARATION 

I, ALADELE, ANDREW KOLAWOLE (22PCO02401), declare that I carried out this 

research under the supervision of Prof. Conrad A. Omonhinmin and Prof. Michael T. Abberton 

of the Department of Biological Sciences, College of Science and Technology, Covenant 

University, Ota, Nigeria and International Institute of Tropical Agriculture, Ibadan, Nigeria, 

respectively. I attest that the thesis has not been presented wholly or partially for the award of 

any degree elsewhere. All sources of data and scholarly information used in this dissertation 

are duly acknowledged. 

 

 

 

 

 

 

 

 

 

ALADELE, ANDREW KOLAWOLE  

 Signature and Date 



v  

CERTIFICATION 

We certify that this thesis titled “MOLECULAR DYNAMICS OF TUBER AND SEED YIELD 

IN Sphenostylis stenocarpa (Hochst. ex A. Rich. Harms) (AFRICAN YAM BEAN)” is an original 

work carried out by ALADELE, ANDREW KOLAWOLE (22PCO02401), in the 

Department of Biological Sciences, College of Science and Technology, Covenant University, 

Ota, Ogun State, Nigeria under the supervision of Prof. Conrad A. Omonhinmin and Prof. 

Michael T. Abberton. We have examined and found this work is acceptable as part of the 

requirements for the award of Masters of Science (M.Sc.) degree in Biology. 

 

 

Prof. Conrad A. Omonhinmin  

(Supervisor)                                                                                             Signature and Date 

 

 

 

Dr. Isaac O. Ayanda  

(Head of Department)                                                                        Signature and Date 

 

 

 

 

 

Prof. Adegoke E. Adegbite  

 (External Examiner)                                                                         Signature and Date 

 

 

 

 

 

Prof. Conrad A. Omonhinmin  

(Dean, School of Postgraduate Studies)                                           Signature and Date



vi  

DEDICATION 

This research work is dedicated to the Almighty God and my parents, Dr. Sunday Aladele and 

Mrs. Grace Aladele 



vii  

AKNOWLEDGEMENTS 

I want to appreciate the most-high God, the custodian of all wisdom, knowledge, and 

understanding, for His grace and favor throughout the duration of carrying out my M.Sc. 

programme. I acknowledge the University Vice Chancellor, Prof. Timothy A. Anake; the 

Registrar Mr. Emmanuel Igban; the Dean School of Postgraduate Studies, Prof. Conrad A. 

Omonhinmin; The Sub-Dean School of Postgraduate Studies, Dr. Hezekiah O. Falola; the 

Dean, College of Science and Technology, Prof. Adewunmi Adedapo Oluwatayo and the Dean, 

Student Affairs, Mrs. Olushola E. Coker. 

I would like to acknowledge the Head of Department, Prof. Isaac O. Ayanda for his 

contribution during my programme. My sincere appreciation goes to my supervisor Prof. 

Conrad A. Omonhinmin and co-Supervisors Prof. Michael T. Abberton, Dr. Olaniyi Oyatomi, 

Dr Rajneesh Paliwal for their support and mentorship given to me all through the stages of my 

research work, I would like to appreciate the Head of Unit, Dr. Samuel A. Ejoh; the 

Postgraduate coordinator, Dr Patrick O. Isibor; My lecturers, Prof. Oluwole O. Obembe, Dr. 

Bello O. Adetutu, Dr. Patrick O. Isibor and the entire staff of the Department of Biological 

sciences for their support. 

I would like to express my profound gratitude to Crop Trust in conjunction with GRC, IITA 

for sponsoring my M.Sc. programme. I also want to appreciate Dr. Olaniyi A. Oyatomi and Dr. 

Rajneesh Paliwal for their contributions and supervision during my research work. I appreciate 

staff and colleague of the Genetic Resource Centre, International Institute of Tropical 

Agriculture. My sincere appreciation goes to my colleagues: Miss Tobi Obadire, Mr. Daniel 

Balogun, and Mr Samuel Oyegbade to name a few whose contributions were invaluable 

through my postgraduate studies. God bless and reward you abundantly. 

I would like to appreciate my supportive parent (Dr. Sunday E. Aladele and Mrs. Grace 

Aladele) and my brother (Victor O. Aladele) for their continuous support. 



viii  

TABLE OF CONTENTS 
CONTENTS PAGES 

COVER PAGE i 

TITLE PAGE ii 

ACCEPTANCE iii 

DECLARATION iv 

CERTIFICATION v 

DEDICATION vi 

ACKNOWLEDGEMENTS vii 

TABLE OF CONTENTS viii 

LIST OF TABLES xi 

LIST OF FIGURES xii 

LIST OF PLATES xii 

LIST OF ABBREVATIONS xiv 

ABSTRACT xvi 

 

CHAPTER ONE: INTRODUCTION          1 

1.1 Background to the Study                                                  1 

1.2 Research Questions          5 

Aim and Objectives:          5 

1.3 Justification for the Study           6 

1.4 Scope of Study          6 

CHAPTER TWO: LITERATURE REVIEW           7 

2.1 Legumes          7 

2.2 Root Tuber Crops          8 

2.3 Tuberous Legumes          9 

2.4 Legumes in Africa         10 

2.5 Germplasm Conservation         13 

2.6 Agronomy and Botany of Sphenpstylis stenocarpa         14 

2.7 Eco-geographical Distribution of Sphenostylis stenocarpa        15 

2.8 Economic Potentials and Uses of Sphenostylis stenocarpa        15 



ix  

2.9 Limitations of Sphenostylis stenocarpa       18 

2.10 Genetic Diversity within Species         18 

2.11 Morphological Characterization of Sphenostylis stenocarpa        19 

2.12 Molecular Characterization of Sphenostylis stenocarpa        22 

2.13 Concept of Yield Stability                                   23 

2.14 Quantitative traits and Marker-Trait Association         24 

2.15 Gene and protein relationship         25 

 

CHAPTER THREE: MATERIALS AND METHODS         27 

3.1 Research site         27 

3.2 Germplasm used         27 

3.3 Experimental Design and Field Plot Management         27 

3.4 Collection of Data         28 

3.5 Equipment         33 

3.6 Reagents         34 

3.7 Methods         35 

 

CHAPTER FOUR: RESULTS          37 

4.1 Phenotype Analysis         37 

4.2 Genome-wide Association Studies (GWAS)         57 

CHAPTER FIVE: DISCUSSION          71 

GWAS         72 

 

 

 

 



x  

CHAPTER SIX: CONCLUSION AND RECOMMENDATION       73 

6.1 Conclusion        73 

6.2 Contribution to Knowledge        73 

6.3 Recommendations        74 

6.4 Limitations to this Study        74 

 

REFERENCES       75 



xi  

LIST OF TABLES 

TABLES TITLE OF TABLES PAGES 

1.1: Research gaps 26 

3.1 : AYB passport information displaying the nation of origin, the acquisition date 29 

4.1 : Seed raw data of the accessions and their replicates 38 

4.2 : Tuber raw data transformed into categorized data 50 

4.3 : Total number of filtered significant SNPs 58 

4.4 :   BLUE FarmCPU Seed thickness 63 

4.5 :   BLUE FarmCPU Tuber weight Group 64 

BLUE FarmCPU Tuber weight 65 

BLUE GLM Tuber weight Group 66 

BLUE GLM Tuber weight 67 

BLUE MLM Tuber weight 68 

BLUE BLINK Pod width 69 

BLUE BLIK Seed thickness 70 4.9: 

4.8: 

4.7: 

4.6: 

4.5: 

4.4: 



xii  

LIST OF FIGURES 

FIGURES TITLE OF FIGURES PAGES 

2.1 :  Diverse tubers and seed of African yam bean 17 

4.1 :  The cladogram showing the clustering pattern of the accessions based 46 

on seed yield 

4.2 : Biplot analysis of seed yield performance and their influencing traits 47 

4.3 : Identification of correlation of traits on the tuber yield 48 

4.4 : The cladogram showing the clustering pattern of the tuber-producing accessions 53 

4.5 : Biplot analysis of tuber yield performance and their influencing traits 54 

4.6 : Identification of correlation of traits on the tuber yield 55 

4.7 : Identification of the relationship between seed and tuber traits 56 

4.8 : FarmCPU model showing significant SNPs for the BLUEs Tuber weight 59 

utilizing Manhattan plot and Q-Q plot 

4.9 :  BLINK model showing significant SNPs for the BLUEs Tuber weight 60 

utilizing Manhattan plot and Q-Q plot 

4.10 :  BLINK model showing significant SNPs for the BLUEs Seed Thickness 61 

utilizing Manhattan plot and Q-Q plot 

4.11 : BLINK model showing significant SNPs for the BLUEs Pod Width utilizing 62 

Manhattan plot and Q-Q plot 



xiii  

LIST OF PLATES 

PLATES TITLE OF PLATES PAGES 

 

 

2.1 : African yam bean staked on-field. Green matured pod in view 22 



xiv  

LIST OF ABBREVIATIONS 

AFLP: Amplified Fragment Length Polymorphism 

AYB: African Yam Bean 

Bp: Base pairs 

 

BLUE: Best linear and unbiased estimators 

Chr: Chromosome 

CTAB: Cetyl trimethylammonium bromide 

DArT: Diversity array technology 

DArTseq: Diversity array technology sequence (DArTseq) 

DNA: Deoxyribonucleic Acid 

EDTA: Ethylene Diaminetetraacetic Acid 

FAO: Food and Agriculture Organization 

GLM: General linear Model 

GRC: Genetic Resource Centre 

 

GWAS: Genome-Wide Association Study 

 

IITA: International Institute of Tropical Agriculture 

ISSR: Inter-Simple Sequence Repeat 

MAF: Minor allele frequency 

MAS: Marker assisted selection 



xv  

MLM: Mixed linear model 

 

mRNA: Messenger ribonucleic acid 

NA: Not available 

NGS: Next generation sequencing 

PCA: Principal component analysis 

PCR: Polymerase chain reaction 

Q-Q: Quantile-quantile 

QTLs: Quantitative Trait Loci 

RAPD: Random Amplified Polymorphic DNA 

SNPs: Single Nucleotide Polymorphisms 

SSR: Single Sequence Repeat 

TASSEL: Trait Analysis by Association, Evolution and Linkage 

tRNA: Transfer ribonucleic acid 

TSs: Tropical Sphenostylis stenocarpa 



xvi  

ABSTRACT 

Sphenostylis stenocarpa (Hochst. ex A. Rich. Harms) (African yam bean - AYB) is an 

understudied and opportunity crop with the potential to contribute to food security It is a 

versatile legume that produces both edible seeds and tubers. AYB faces under-exploitation due 

to limited understanding and challenges as well as an unabetted threat to its diversity like 

several other indigenous plant species. Expanding the diversity of global food sources has 

become imperative. This study investigated the molecular dynamics in AYB by assessing the 

genetic diversity in non-tuber and tuber-producing landraces, identified SNP markers 

associated with tuber formation and determined the relationship between seed and tuber 

production. The study evaluated accessions from the Genetic Resources Centre of the 

International Institute of Tropical Agriculture (IITA), Ibadan. Phenotypic data were generated 

from monitored growth, genotyping was conducted using DArTseq technology, with SNP data 

generated afterward. Phenotypic data employed clustering, correlation analysis and distribution 

plots. Genome-wide Association Studies (GWAS) using the GAPIT package in R elucidated 

population structure and identified SNPs responsible for yield. Multiple traits such as Number 

of Pods, Hundred Seed Count, Total Seed Weight per Plant, Total Pod weight indicate a close 

association with each other showing a strong indication with the accessions and yield. While 

clustering for seeds and tubers showed four and five clusters respectively. BLUEd traits and 

2254 SNP markers from 92 genotypes were used for the association analysis. Using the 

BLINK, FarmCPU, GLM, and MLM models. Twelve significant SNP markers were identified 

to be associated with three African yam bean yield traits (Tuber weight, Seed thickness, and 

Pod width). These results have the potential to accelerate marker-assisted selection in 

molecular breeding. 

 

 

 

Keywords: African yam bean, Opportunity, Seed, Tuber, SNPs, Yield 



1  

CHAPTER ONE 

INTRODUCTION 

1.1 Background to the Study 

 

The genus Sphenostylis E. Meyer (Leguminosae; Papilionoideae; Phaseoleae) has seven species, 

including the African yam bean (AYB), which grows in dry woods and open or wooded savannas 

throughout tropical and Southern Africa (Oagile, Davey & Alderson, 2007). The hard-to-cook 

African yam bean (AYB) is a leguminous plant that is underutilized and is widely produced in 

Western Africa (Uchegbu, 2015), Eastern Africa, and Central Africa (Bhat & Karim, 2009). When 

grown at multi-locations, AYB is primarily used as a staple crop and is considered a security crop 

for farmlands that are fallow and getting ready for a new planting season. Growing for subsistence, 

local farmers typically plant this perennial crop (George, Obilana & Oyeyinka, 2020), (Enujiugha 

et al., 2012). When the crop is harvested, it yields consumable seeds and tubers that are mosly 

eaten in South-eastern Nigeria (Idowu, 2014) and other parts of West Africa (Ajibola & Olapade, 

2016). The crop exhibits a high degree of genetic variety (George, Obilana, & Oyeyinka, 2020), 

and recent research has looked into new accessions (Baiyeri et al., 2018). 

The Sphenostylis stenocarpa (Hochst. ex A. Rich. Harms) (AYB) is the most economically 

significant species among the seven species that are currently recognized in the genus Sphenostylis 

(Potter, 1992). A few farmers who had relied on this native crop—whose pods and tubers are 

edible—appreciate AYB very much because it was an inexpensive source of protein. Despite being 

a crop with good nutritional potential and growing in a well-adapted habitat, the African yam bean 

is nevertheless considered an underutilized and neglected plant (Aina et al., 2021). 

AYB has enormous economic potential, and its seed and tuber contain a higher protein content 

than most other tuberous legumes (Dakora, 2013). In livestock feed formulation, grains are an 

equal or superior replacement for common legumes, and their medicinal properties to treat certain 

terminal diseases are worth investigating. Both humans and animals can safely consume seed and 

tubers, which provide significant nutritional advantages (Adewale & Nnamani, 2022). The crop 

tolerates a wide range of climatic, geographical and edaphic ecologies (Potter & Doyle, 1992). It 



2  

can flourish in shady spots where other pulses are unable to thrive. (Okpara & Omaliko, 1995). As 

a native crop, AYB is mostly grown in a mixture of Dioscorea spp, Manihot esculenta, Zea mays, 

and Sorghum bicolor. Mostly, Manihot esculenta, Zea mayz, and Sorghum bicolor serve as stakes 

for yam and also support for the crop (Yusuf et al., 2022). Africa, if cultivated extensively. AYB 

is recommended in ethnomedicine to treat high blood pressure and diabetes. In the Nigerian state 

of Benue, it is used to treat mumps. It implies that certain phytochemicals, minerals, and 

antioxidants found in African yam beans may have advantageous physiological and/or 

pharmacological benefits (Baiyeri et al., 2018). 

Agu (2008) and Ameh & Okezie (2006) have found pests and illnesses that prey on field-planted 

AYB. Based on the crop's developmental stages, the pests that were found can be divided into two 

groups: pests at the stage of vegetation and pests at the reproductive stage. During the vegetative 

stage of AYB, the following organisms can attack: Maruca testulalis, Cydia ptychora, 

grasshoppers (Zonocerus variegatus), cutworm larvae and/or adults (Agrotis spp.), and leaf- 

rolling caterpillars (Sylepta derogate). Cydia ptychora, Heliothis armigera, Riptortus dentipes, 

Apion varium, and Nezara viridula were discovered on the crop during the reproductive stage 

(Ameh & Okezie, 2006). Pathogens that cause disease in AYB include Aecidium spp., Phoma spp., 

and Oidium spp. They have been found to be potent pathogens that cause stem rust, leaf spot, and 

powdery mildew, in that order. Diseases in AYB that have been identified are wilting leaf mosaic 

and root gall (Ameh & Okezie, 2006; Agu, 2008). Additionally, a 1999 study by Omitogun et al. 

revealed that adding crude lectin-enriched extracts from Sphenostylis stenocarpa (Harms), the 

African yam bean, to artificial seeds (5.0%) significantly impacted the nymphs' survival (Okeola 

& Machuka, 2001). 

The species is used extensively in sociocultural contexts. It has been utilised as an alternative 

medicine to heal certain illnesses as well as for a variety of cultural rituals, such as weddings and 

festivals. Because of the anti-nutritional components of the crop and the length of time needed to 

cook the grains, it is still an underutilized grain legume (Ojuederie et al., 2021). Humans are not 

able to digest complex carbohydrates like oligosaccharides, which are present in African yam 

beans (AYB). It is recognized that oligosaccharides are prebiotics or food for probiotics. Therefore, 

adding prebiotic supplements to milk helps yoghurt starts thrive. (A. Aderinola & Y. Olanrewaju, 

2014). 



3  

A concerted effort has been made to prevent the extinction of African yam bean species by 

diversifying their uses and encouraging their use. A few fermented foods made from AYB contains 

a product that resembles vegetable milk, and yoghurt (Amakoromo et al., 2012). Samples of cow's 

milk yoghurt that were replaced with AYB milk—made from dehulled seeds and kept at 40 degrees 

Celsius for up to 14 days—had good quality, particularly the 50:50 (African yam bean milk: Skim 

milk) ratio (A. Aderinola & Y. Olanrewaju, 2014). A small amount of AYB germplasm is also kept 

in the Genetic Resources Centre at the International Institute of Tropical Agriculture (IITA), 

located in Ibadan, Nigeria. Poor conservation technology has several drawbacks, such as limiting 

reach to genotypes for farmers, scientists, and breeders; making AYB vulnerable to natural disasters; 

causing genetic erosion; and reducing crop diversity (Nnamani et al., 2021). 

Previous research have exploited the diversity of physical features found in AYB to divide the 

different accessions into discrete groups according to phenotypic data (Akande, 2009; Ojuederie 

et al., 2015). Four to six clusters of genotypes were identified based on an analysis of the 

intraspecific variability in AYB accessions' phenotypic and genotypic characteristics (Ojuederie 

et al., 2015; Nnamani et al., 2019). Attempts to characterize some of the available AYB accessions 

using a morphological approach have been carried out by (Ibirinde et al., 2019; Ibirinde et al., 

2019; Aina et al., 2020). This has helped in the provision of valuable distinguishing traits that are 

currently being used to distinguish accessions of AYB. 

Researchers have also utilized PCR-based markers to characterize the variability in the genomes 

of AYB. Polymerase Chain Reaction based markers e.g Amplified Fragment Length Polymorphic, 

RAPD, Simple Sequence Repeat, and ISSR have previously been used in conducting genetic 

diversity studies on AYB germplasms (Adewale et al., 2014; Moyib et al., 2008; Popoola et al., 

2011; Omena et al., 2014; Shitta et al., 2016). For research on genetic diversity, more exact 

molecular markers have also been employed, such as SNP markers (Olomitutu et al., 2022). Other 

Next-generation molecular markers (such as Diversity Array Technology) are currently being used 

to explore the diversity among GRC-IITA AYB germplasm (Paliwa et al., 2020). However, 

Adewale et al. (2015) suggest a shift in focus to trait-marker association studies in AYB 

germplasm. In any crop, yield is a major agronomic factor or concern for individuals interested in 

propagating or breeding. The emphasis on solving global hunger will only be attained if yield 

participates in whatever strategy is set in place or being considered. Because yield is primarily 



4  

determined by genotype, environment, and crop management techniques, Sphenostylis stenocarpa 

grain yields in Nigeria are modest and vary by regions: 205–631 kg ha-1 in Abakaliki (Ogah, 

2013), 440 kg ha-1 in Mokwa, and 749 kg ha-1 in Ibadan (Adewale, Ojo, & Abberton, 2017). The 

high expense of obtaining the stakes required for good yields in Southeast Nigeria has further 

reduced production, and only a small number of local farmers are able to grow the crop in 

accordance with their indigenuous cropping practices (Baiyeri et al., 2018). More research will 

need to be conducted on the yield of specific accessions that show promising yields. 

More than two billion people rely on tuber and root crops for their energy and nourishment, and 

they are a significant source of revenue in rural and marginalized areas. They are being utilized as 

feed for livestock and as crude substances for industrial processes, in addition to their many 

functions as cash crops, regular food crops, and security crops (Scott et al., 2000). Since the tubers 

and seeds provide food for both people and animals, they are commercially significant. Its grains 

and tubers contain roughly 29% and 19% crude protein, respectively, making it a good source of 

protein (Aki̇nyosoye, 2022). A lot of time is spent processing and cooking AYB because of the 

anti-nutritional ingredients it contains. Typically, it is boiled and consumed by itself or with rice, 

yam and corn. When preparing food, it can take the place of cowpeas, particularly in the lean 

season when food is in short supply for rural farmers (Akande, 2009). 

Since indigenous legumes are becoming more and more important for food security in Africa, 

achieving yield stability despite increased from one season to season yield variance is an important 

breeding goal (Thiyagu et al., 2012.). Sustaining yield stability despite season to season variability 

is essential to the productivity of the AYB. Determining how steady and consistent the production 

of such crops requires investigating crop yield stability (Aremu et al., 2020). The potential for 

understanding the basis of genetic variation in complicated traits at the genome-wide level has 

been made possible by the current rapid advancements in omics technology. 

One excellent example of high throughput technology is Diversity Array Technology (DArT), 

which has high call rates, high repeatability, and a comparatively low cost. Most notably, DArT 

can provide genome-wide SNPs in species like AYB that have no previous DNA sequence data 

(Edet et al., 2018; Barilli et al., 2018). Several legumes have benefited from the use of the DArT 

technique in genetic investigations, including soybeans (Vu et al., 2015), common beans 



5  

(Valdisser et al., 2017), and pigeon peas (Yang et al., 2011). Originally created for gene mapping 

of human diseases, Association Mapping (AM) (Olomitutu et al., 2022), is currently a widely used 

technique in research of plant maker-trait associations. Association mapping uses recombination 

events that accumulate over generations in natural populations to identify linkage disequilibrium 

between genes and genetic markers governing the characters of interest (Ruggieri et al., 2014). It 

assesses if a population's distribution of specific alleles with particular traits is more common than 

predicted (Flint-Garcia et al., 2005). Putative QTLs and genes in numerous leguminous crops have 

been found through GWAS reports (Gali et al., 2019; Ahmed et al., 2021; Rajendran et al., 2021). 

These findings have been crucial in helping to understand the inheritance of quantitative traits 

(Goddard et al., 2016) and the deployment of traits through marker-assisted selection (Uwera et 

al., 2021). Several Quantitative Trait Loci (QTLs) for seed size features in common beans (Mir et 

al., 2021; Giordani et al., 2022), soybeans (Li et al., 2019), and cowpeas (Lo et al., 2019) have 

been found in legumes. 

 

1.2 Research Questions 

 

I. What Single Nucleotide Polymorphism (SNPs) are responsible for tuber formation 

 

II. Does seed yield influence tuber yield 

 

III. What is the level of diversity present in tuber-producing available accessions? 

 

Aim and Objectives: 

This research aimed to determine the genetics of tuber and seed yield in African yam bean. 

The specific objectives of this study, therefore, were to: 

I.  assess the genetic variances for tubers by determining the extent of genetic variability 

present within the population regarding tuber-related traits. 

II. determine the relationship between seed yield and tuber yield in the studied accessions. 

III. identify SNPs associated with tuber formation. 



6  

As a legume with huge biomass and fixes nitrogen. African yam bean therefore, has the potential 

of enhancing soil organic matter through biological nitrogen fixation and improving soil fertility 

through the decomposition of leaves and other plant parts. In spite of the huge potentials of the 

crop, it is faced with some constraints that have limited its production and led to its neglect and 

underutilization. Some of these constraints are: high cost of production arising fromstaking its vine 

(staking is a necessity for good yield), genetic erosion of African yam bean biodiversity in major 

growing areas especially in Nigeria and Ghana, lack of improved varieties (Baiyeri, et al., 2018). 

and information on the agronomic management practices of the crop is very scanty in the literature. 

 

1.3 Justification for the Study 

 

The instability of tuber-producing accession is a major concern for farmers and the breeding 

community targeting African yam bean. The need to ascertain genes responsible for tuber 

formation will provide researchers with basis for further study in genome editing and gene 

engineering for stable or desirable tuber yield. To date, no genome-wide study has been conducted 

on the SNPs responsible for tuber formation traits. The genome-wide study conducted on SNPs 

responsible for seed traits was conducted by (Olomitutu et al., 2022). Understanding the SNPs 

responsible for tuber formation will help speed up breeding exercises. 

 

1.4 Scope of Study 

 

A total of one hundred African yam bean accessions were collected from the Genetic Resource 

Centre (GRC), Internation Institute of Tropical Agriculture (IITA), Ibadan. The scope of this 

research was to identify significant SNPs that may be associated with the AYB tuber and seed 

yield. The GWAS was based on data for 2023/2024 planting seasons. Missing data were estimated 

using BLUEs. 



7  

CHAPTER TWO 

LITERATURE REVIEW 

2.1 Legumes 

Fabaceae or Leguminosae is the family that includes commercially significant legumes, such as 

grains, oilseed crops, fodder crops, shrubs, and tropical or subtropical trees. Both people and 

animals may obtain high-quality protein from legumes. By producing their own nitrogen in 

conjunction with nitrogen-fixing bacteria, they help improve the soil. After daisies (Asteraceae or 

Compositae) and orchids (Orchdaceae), they belong to the third-largest family of plants that bloom 

(Singh, Chung & Nelson, 2007). Approximately 9.4% of all flowering plant species are found in 

the biggest genera, which include Astragalus (with over 2,400 species), Acacia (with over 950 

species), Indigofera (about 700 species), Crotalaria (about 700 species), and Mimosa (about 500 

species) (Magallon & Sanderson, 2001). 

 

Leguminous, from the Latin legumen, meaning "seeds collected in pods," is where the name 

legume originates. Pulses are the name given to legumes, especially low-fat varieties, in 

Bangladesh, India, Canada, and other parts of the world. Legumes, or pulses, have been a staple 

diet for farmers since the Neolithic Revolution, when human farming methods first appeared. A 

collection of "founder crops" known as the "Big Eight" were domesticated in the Fertile Crescent 

between the tenth and ninth millennia BCE. Asouti and Fuller (2013) list the legume (Lens 

culinaris), chickpea (Cicer arietinum), pea (Pisum sativum), and bitter vetch (Vicia ervilia). Over 

Eight hundred and twenty million people suffer from inadequate food, while a large number of 

others have poor diets that result in shortages in some micronutrients (Webb et al., 2018). 

 

Through atmospheric nitrogen fixation in root nodules in symbiosis with soil bacteria from the 

families Bradyrhizobium, Rhizobium, and others, they are self-supporters of nitrogen fertilization. 

After extensive research into the genetic regulation of these processes, almost two-hundred genes 

necessary for legumes' symbiotic-nitrogen fixation have been found using a variety of forward- 

and reverse-genetic techniques (Roy et al., 2020). 



8  

2.2 Root Tuber Crops 

A recent analysis from the Consultative Group on International Agricultural Research (CGIAR), 

revealed that "by 2020, root crops will be many things to many people" (Scott and others, 2000). 

Root crops generate their edible energy-rich underground plant structures from modified roots, in 

contrast to tuber crops, which have edible energy-rich storage organs that partially or entirely 

emerge from subterranean stems (Okuneye, 2001). They are grown year-round in Africa's sub- 

Saharan region. The most significant food crops for direct human consumption in Africa are the 

tuber crops (yam, cocoyam, and sweet potato) and the two primary root crops—cassava and sweet 

potatoes. These crops, which account for about 240 million tonnes yearly and are farmed on over 

23 million hectares, are cultivated in various agro-ecologies and production techniques. 

 

All other staples in Africa are surpassed in value by yam, cassava, potatoes, and sweet potatoes 

combined, even cereal crops (which are grown yearly on 108 million hectares of land and yield an 

average of 169 million tonnes of grains) (Nanbol & Namo, 2019). AIn line with these worldwide 

developments, growers of RTBs like yam, cassava and potatoes grew more focused on the market, 

placing an increased emphasis on cash sales to supplement their more conventional function as 

crops for food security. In poor nations, informal cross-border and domestic trade in RTBs is 

widespread. All of the major food commodities together have not produced as much cassava as 

SSA has during the past 30 years, despite SSA's cassava output growing at a faster rate. Sixty- 

seven percent of the increase in cassava output was focused in Nigeria, the Democratic Republic 

of the Congo (DRC), and Ghana, where it is the main staple or co-staple (Scott, 2021). 

 

Two different strategies were used in the robust growth of the potato crop, which was grown in 40 

countries throughout the region. Algeria, Morocco, Egypt, and South Africa are nations that 

practice irrigation agriculture. As an alternative, rain-fed potato farming is practiced in SSA 

nations like Tanzania and Nigeria. In SSA, sweet potatoes evolved in a very different way. 

Harvested area rose via 3.9 million hectares, while yield climbed via 22.4 million tonnes. Out of 

eleven primary food items available in the area, the average growth rates for production from 1988–

1990 



9  

to 2016–18 were greater (5.4% yr-1) than any other, and the area harvested grew at a rate of 4.5% 

annually, only surpassed by yams (Scott, 2021). 

 

While there are over 600 species of yam, some native to Asia, more travelers head to the Caribbean, 

and some to West Africa (Asiedu & Sartie, 2010), The countries of Ghana and Côte d'Ivoire, 

Nigeria, accounted for 88% of that growth. Of the 24 SSA nations that cultivate yam, Ghana, 

Nigeria, and Côte d'Ivoire have harvested 90% of the region's total production from 1961–1963, 

with Nigeria producing 68% of the crop overall (Scott, 2021). In West Africa, yam is valued for 

its use as a commercial crop and staple in rural areas, a significant part of traditional cultural and 

ceremonial events, a secondary urban diet, a source of foreign currency, and more (Nweke, 2016; 

Frossard et al., 2017). 

 

2.3 Tuberous Legumes 

Many names for Vigna vexillata (L.) A. Rich (subgenus Plectrotropis; family Fabaceae) exist, 

including wild cowpea, tuber cowpea, zombi pea, and so forth. It is a promising and underutilized 

legume. This herbaceous legume is pan-tropical and can be found on the continents of America, 

Asia, Australia, and Africa. According to Tripathi et al. (2021), it is one of the underutilized 

legumes with the potential to be exploited commercially. Pachyrhizus tuberosus is a type of 

tuberous legume that is occasionally cultivated in South America's tropical lowlands. In his 

Chartas ineditas (1556), Padre Jose de Anchieta most likely described P. tuberosus and its 

applications from the island of Saño Vicente, near the port of Santos, Brazil. The largest species 

in the genus, this herbaceous vine can reach lengths of up to +10 m on its stems (Sørensen et al., 

1997). 

 

Apios (Apios americana Medik.), commonly known as the "potato bean," is a nitrogen-fixing 

legume that grows underground stolons to produce tubers rich in protein at nodes. It is native to 

eastern North America. Apes were the primary source of food for indigenous people on the east 

coast of North America, and they may be grown as a crop (Belamkar et al., 2015). One description 

of the flavor of tubers is a combination of Irish potatoes (Solanum tuberosum L.) and boiled 

peanuts (Arachis hypogaea L.). Tubers make great chips because they contain few quantities of 

reducing sugars (Carlisi & Wollard, 2005). Because apios has more nutritional value than potatoes, 



10  

sweet potatoes (Ipomoea batatas (L.) Lam.) and taro (Colocasia esculenta (L.) Schott), it is being 

cultivated and consumed as a food crop in South Korea and Japan (Kikuta et al., 2011). 

 

Just one leguminous species, the winged bean, has the capacity to produce both tuber and pod yield 

simultaneously. The tuber's nutrients and protein value make it a viable crop substitute for animal 

feed (Rakvong et al., 2024). Often referred to as the "winged bean," Psophocarpus tetragonolobus 

(L.) DC. is an evergreen tropical legume crop which is a member of the Papilionoideae subfamily 

and the Fabaceae family. According to Vantanparast et al. (2016), the size of its diploid genome, 

with nine chromosomal pairs in it. (2n = 2× = 18), is estimated to be 1.22 gigabase pairs. Southeast 

Asian nations have long been consumers of winged bean tubers. As such, they are considered 

staple foods in several regions of Myanmar and Indonesia (Tanzi et al., 2019). In Papua New 

Guinea, the tubers can be cooked or grilled in banana leaves to make a variety of dishes. According 

to Vantaprast et al. (2016), It can also be cooked and added to tempeh or other traditional 

Indonesian recipes. 

 

2.4 Legumes in Africa 

The most prevalent family in dry forests and tropical rainforests in America and Africa is the 

Fabaceae (Burnham & Johnson, 2004). The tropics are home to more than thirty species of 

economically significant legumes (Raemaekers 2001; Gowda et al 2007). Among them are the 

soybean (Glycine max), cowpea (Vigna unguiculata), pigeon pea (Cajanus cajan), groundnut 

(Arachis hypogaea), chickpea (Cicer arietinum), and common bean (Phaseolus vulgaris). Field 

pea (Pisum sativum), faba bean (Vicia faba), lentil (Lens culinaris), hyacinth bean (Lablab 

purpurea, also known as Dolichos lablab), Kersting's groundnut (Macrotyloma geocarpum), lima 

bean (Phaseolus lunatus), African yam bean (Sphenostylis stenocarpa), mung bean or green 

gramme (Vigna radiata), black gramme or black bean (Vigna mungo) and moth bean (Vigna 

aconitifolia) are some other noteworthy legumes that grow. With over 70 species from Central and 

North America, Phaseolus is a broad and varied genus of pulse crops (Bellucci et al., 2014). Five 

of these species—P. vulgaris, P. dumosus, P. coccineus, P. acutifolius, and P. lunatus—have 

reached domestication, while a few more—Delgado-Salinas, show indications of approaching 

domestication (Bibler & Lavin, 2006). 



11  

Phaseolus vulgaris L., often referred to as the common bean, it is the most significant edible 

legume globally for direct use, producing close to 12 million metric tonnes annually. Three- 

quarters of this crop is farmed in sub-Saharan Africa and Latin America, which are its main 

producing locations (Akibode & Akibode, 2011). The majority of the Phaseolus species are 

thought to be Mesoamerican in origin based on their geographic distribution (Delgado-Salinas, 

Bibler & Lavin, 2006). According to Bennett and Leitch (2011), With 22 chromosomes and a 

haploid genome estimated to be between 587 and 637 megabases in size, P. vulgaris is a true 

autogamous diploid species. 

 

Grown throughout the semi-arid sub-Saharan region of Africa, the African legume known as 

"Bambara groundnut" (Vigna subterranea (L.) Verdc) is called such (Hillocks, Bennett & Mponda, 

2012). Due to its hardiness, this crop has been acknowledged as a valuable and nutrient-dense 

source of food during times of scarcity (Mbosso et al., 2020). Its climate-smart characteristics, 

such as its capacity to fix nitrogen and thrive in unfavorable environments like poor soils and 

drought, may be to blame for this (Paliwal et al., 2020; Mayes et al., 2019). Legumes such as Vigna 

subterranea (L.) Verdc, or bambara groundnut can supplement the diets of households 

experiencing food insecurity by offering vital nutrients. According to Mazahib et al. (2013), the 

seed has 65% carbohydrates, 18% protein, 6.5% oil, and vital minerals like calcium, iron, 

potassium, and sodium. 

 

Bambara groundnut has a number of ANFs that have been found, just like other legumes. The 

digestion and bioavailability of vital nutrients may be adversely impacted by their presence. ANFs 

found in Bambara groundnut that are frequently reported include phytic acid, condensed tannins, 

and trypsin inhibitors. The testa is home to condensed tannins, which are more prevalent in seeds 

with deeper colors (Nti, 2009). Although these polyphenolic compounds have antioxidant 

properties, they can combine with carbohydrates, proteins, and dietary minerals to produce 

indigestible complexes that lower their bioavailability ((Mohan, Tresina & Daffodil, 2016, Unigwe 

et al., 2018). 

 

One of the main obstacles to the utilization of Bambara groundnut is its hard-to-cook (HTC) 

feature (Mubaiwa et al., 2017). According to Tan et al. (2020), the HTC trait is linked to legume 



12  

cotyledon resistance to softening during cooking, necessitating a longer cooking period to achieve 

the desired texture. When still young and immature, Raw or roasted Bambara groundnut grain is 

used as a snack. The grain must be boiled before being processed or eaten because of its tough 

seed covering as it ages. Grain is ground into flour in Benin and used to produce dumplings, cakes, 

and bread. Soup is made from crushed grains in the countries of eastern Africa. Furthermore, in 

Zambia, bread is made using processed Bambara groundnut flour (Majola, Gerrano, & Shimelis, 

2021). 

 

The grains are used to flavor cooked cowpea grain in Swaziland and South Africa (Mubaiwa et 

al., 2017). It is also processed to make milk that resembles soybean milk. Many African nations 

utilize the resulting milk as a weaning meal. When compared to soybean milk, Bambara groundnut 

milk is lighter in color (Yao et al., 2015). For animal feed, the biomass—the leaves, stem, and 

haulm—has been utilized extensively. While the grains are fed to pigs and poultry, the seed cake 

is also utilized as animal feed. Young, succulent leaves are rich in vital mineral elements that are 

beneficial to animal health, such as phosphorus and nitrogen (Keller, 2004). Bambara groundnut 

leaf protein is given to tilapia fish in Nigeria (Mazahib et al., 2013). 

 

Because legumes fix atmospheric N2, they make inexpensive, environmentally friendly N 

fertilizers. Legumes are also included in cycles based on oilseed rape and cereals as diversification 

crops. Despite these advantages, less than 4% of Europe's arable land is used for legume crops, 

and the continent's legume seeds are underutilized for feeding both humans and animals. However, 

for the sake of sustainable agriculture, European authorities are currently encouraging the growth 

of legume crops (Voisin et al., 2014). Even with their benefits, legume cultivation has not kept up 

with expectations and is still less than that of other crops like grains. Alongside this, industrialized, 

mostly cereal-based farming practices that primarily rely on fossil fuels have gradually replaced 

traditional farming methods. Consequently, over the past 50 years, the acreage of the majority of 

temperate legumes (pea, faba bean, vetches, and lupin) has decreased globally (Rubiales & Mikic, 

2015). Reduced and unstable yields as well as vulnerability to situations of biotic and abiotic stress 

are additional factors that lead to the low diffusion of legume cultivation; According to Stangnari 

et al. (2017), average yields per unit area have grown (soybean +86%, lentil +77%, groundnut 

+75%, and chickpea +70%) but less than cereal crops (+104%, on average). 



13  

Although they seem to be a great protein source, legumes represent currently only a small portion 

of the diets of most people. Globally, the average daily consumption of meat and legumes is 112 

g/person and 21g/peron, respectively. Due to their low carbon and water footprints, ability to fix 

nitrogen into the soil, affordability, and status as a sustainable source of protein, legumes are a 

staple in most traditional meals across various cultures (Semba et al., 2021). Legumes are capable 

of fixing nitrogen from the air, therefore they do not need to be fertilized with nitrogen. Thus, the 

reduction of nitrogen fertilizer inputs is made possible by their inclusion into cropping systems. 

Their crop residues' mineralization allows for a reduction in nitrogen fertilization of the subsequent 

crop as well (Voisin et al., 2014). Therefore, in order to achieve a comparable or greater yield, 

Carrouée et al. (2012) recommend lowering the application of nitrogen following pea by 20–60 kg 

N/ha on average for wheat, and 30–60 kg N/ha for oilseed rape. The decrease in nitrogen 

fertilization helps to lower greenhouse gas emissions (Jensen & Hauggaard-Nielsen, 2003). These 

emissions include nitrogen protoxide (N2O) released when dispersing and releasing carbon dioxide 

released throughout the Haber-Bosch procedure, which produces fertilizers. Furthermore, N2O 

emission levels from legume crops and their residue mineralization are comparable to those from 

non-legume, unfertilized crops (Jeuffroy et al., 2013). Legumes may therefore have a noteworthy 

effect on limiting the depletion of fossil fuels and lowering the emissions of greenhouse gases, 

which will contribute to reducing climate change (Voisin et al., 2014). 

 

2.5 Germplasm Conservation 

A significant natural resource, germplasm is essential for preserving plant diversity since it can 

reveal information about a species' genetic makeup (Priyanka et al., 2021). Resources for plant 

genetics are the result of combining all allelic sources that affect a crop's spectrum of attributes; 

The genetic material that is transferred from one generation to the next is called germplasm. 

(Priyanka et al., 2021). According to Rao (2003), plausible sources of this genetic diversity include 

landraces or historic varieties, closely similar species of wild plants that are either straightforward 

or indirectly the ancestors of domesticated species, domesticated or semi-domesticated varieties, 

and the cultivars that comprise them no longer still in use or have become outdated. According to 

Baute, Dempewolf, and Rieseberg (2015), mobilizing these allelic resources for efficient and 

sustainable usage is fraught with difficulties despite their availability. 



14  

Due to a lack of long-term maintenance resources, only roughly 30 countries have chosen to save 

their germplasm in gene banks, despite the fact that there are already many gene banks operating 

throughout the world (Kell, Marino & Maxted, 2017). Furthermore, the majority of the In these 

gene banks, 7.5 million accessions belong to crops, including their diverse wild cousins and 

landraces, that are essential to the sustenance of humans and other animals. Nonetheless, certain 

underutilized species and crops of local importance must be preserved (Priyanka et al., 2021). For 

the conservation of diversity in plants, two fundamental conservation tactics in situ and ex situ 

each consisting of a variety of techniques are used (Engelmann, 2012). The preservation of 

ecologies and their natural habitats, as well as upkeep and recovery of healthy groups of various 

species—including domesticated or farmed species—in their native environments, is known as "in 

situ conservation. "While in situ conservation is particularly appropriate for wild species and 

landrace material on farms, ex-situ conservation is beneficial for crops and their wild relatives. 

Engelmann (2012) 

 

2.6 Agronomy and Botany of Sphenpstylis stenocarpa 

African yam bean is a high-protein tuber and seed crop that has the potential to improve food 

security (Adewale et al., 2012). It is a versatile legume that produces both edible seeds and tubers 

(Omonhinmin et al, 2021). Similar to cowpeas, bean seeds are often housed in a pod. Ten to thirty 

seeds, perhaps in multiple colors, are found in each pod. With broad, heart-shaped leaves that are 

spread widely at each node along the stem, the pods are often carried on a climbing stem (Asoiro 

et al., 2011). 

 

In Ghana and Nigeria, AYB planting often begins in May or June, once the rains have stabilized 

(B. Daniel & N. Celestina, 2012). AYB has historically been planted as a secondary crop in mixed 

crops, particularly cassava and yam, in Nigeria and Ghana. It is uncommon to sow AYB as a stand- 

alone crop; instead, it is typically planted alongside yams to share a common support stake 

(Ibeawuchi et al., 2007). Breaking dormancy is not a way to encourage imbibition prior to 

germination in AYB, regardless of how hard the seed is (Olisa, Ajayi & Akande, 2010). Usually 

on the main and subsidiary branches, 4-10 flowers are produced on long peduncles. Its raceme- 

shaped inflorescence shows an acropetal floral maturation mode (Adewale et al., 2012). The 



15  

normal petals of the huge, incredibly gorgeous blooms twist slightly back on themselves at 

anthesis, blending pink and purple. Per the findings of (PopoolaJ. & AgboladeJ., 2011) 

 

The pollen grains exhibited tricolporate, fenestrate, and scabrateexine utilizing 25 AYB 

accessions. Three protuberances (germpores) in a predetermined geometric design broke the 

spinous cover, and the pollen grains are single reticulate, somewhat rounded, and free of sharp 

corners. The flower appears to be self-pollinating; three or more pods are produced on a peduncle. 

When fully developed, the typically long and straight unicarpel pods turn brown. Both sides of the 

pods may have a raised or level edge. The majority of dehydrated pods break and shatter their 

seeds when they dehisce at the ventral and dorsal sutures. On the other hand, different AYB dried 

pod-shattering tendencies were noted by Adewale et al. (2012). Harms (1899) created the botanical 

genus Sphenostylis to encompass a collection of unique leguminous species that were previously 

categorised under the names Dolichos and Vigna. The Greek word sphen, which meaning wedge 

shape, gave rise to the genus name Sphenostylis (Allen & Allen, 1981). 

 

2.7 Eco-geographical Distribution of Sphenostylis stenocarpa 

Sphenostylis stenocarpa can grow in a variety of soil and climate conditions (Aremu et al., 2020). 

African yam beans are classified based on the natural zones in which they have been domesticated; 

their origin cannot be pinpointed. However, there is also the theory that African yam beans 

originated in Ethiopia and then spread to many other tropical African regions (Oagile, Davey & 

Alderson, 2007). Tropical Africa, particularly the nations of Nigeria, Ivory Coast, Ghana, Togo, 

Gabon, Congo, Ethiopia, and other regions of East Africa, is the center of variety for it (Olasoji J. 

O, 2011). AYB is cultivated in Nigeria mainly for seed (Ojuederie & Balogun, 2019). 

 

2.8 Economic Potentials and Uses of Sphenostylis stenocarpa 

West Africa has not paid much attention to the tubers that are consumed in most parts of East and 

Central Africa (Ojuederie & Balogun, 2019). The protein content of AYB is comparable to that of 

common beans, pigeon peas, chickpeas, and Bambara groundnut. In addition, the bean has higher 

or equivalent amounts of dietary fiber (Ndidi et al., 2014; Anya & Ozung, 2019), carbohydrates 

(Ajibola & Olapade, 2016; Nwosu, 2013), and essential minerals (Adamu et al., 2015) like 

magnesium, iron, zinc, and calcium. According to protein fractionation, the two most common 

proteins found in AYB are globulin and albumin (Ajibola et al., 2016). According to Ade- 



16  

Omowaye, Tucker, and Smetanska (2015) and Chinonyerem, Obioha, and Ebere (2017), the bean 

provides the necessary amino acids. 

 

Ojuederie & Balogun (2019) reported that there were generally fewer antinutrients in the tubers 

than in the seeds. They also reported that accessions TSs 107 and TSs 140 were found to have the 

lowest levels of anti-nutrients in harvested tubers and the highest quantities of protein (15.9% and 

15.4%) and carbs (68.7% and 67.9%), respectively. Oluboyega, Olayele, and Ojo (2018) reported 

that some accessions of AYB possess antioxidant-related phytochemicals, such as phenolics and 

flavonoid content, which resulted in appreciable antioxidant activities. This food crop should be 

utilised to produce functional foods because of the presence of these phytochemicals. 

Incorporating AYB into food would not only improve or strengthen it, but also give consumers a 

natural means of maintaining their health (George, Obilana & Oyeyinka, 2020). Germinated 

through its ability to lower blood glucose and avoid hyperinsulinemia, AYB with hypoglycemia 

potential may both directly and indirectly reduce oxidative stress by acting as free radical 

scavengers (Uchegbu, 2015). 



17  

 

Figure 2.1: Diverse tubers and seed of African yam bean. (Source: Adewale & Nnamani, 2022). 



18  

2.9 Limitations of Sphenostylis stenocarpa 

Due to a number of limitations, including lengthy maturation cycles of nine to ten months and 

cooking times of up to twenty-four hours (Njoku, Eli & Ofuya, 1989; Suzzy Shitta et al., 2022), 

AYB potential is still mostly unrealized (Afolabi, Ogunsanya & Lawal, 2019), and the abundance 

of anti-nutrition factors [(Anya & Ozung, 2019; Ndidi et al., 2014; Ajibola & Olapade, 2016; Sam, 

2019), and poor grain yield (Saka et al., 2004) have negatively influenced the adaptability of the 

crop. The distinctive hardness of the seed coat of the AYB is one of its issues (Oshodi et al., 1995). 

This leads to a high energy cost because of the lengthy cooking time, coupled with the agronomic 

need for stakes, and the lengthy maturity period (Okpara and Omaliko, 1995). The seeds contain 

anti-nutrients such as phytic acid, tannins, hydrogen cyanide, trypsin inhibitors, and saponins. 

These substances have the ability to bind with metals and obstruct the body's absorption of 

nutrients and secondary metabolites (NRC, 1979). The African yam bean's sensitivity to 

photoperiod appears to exacerbate the aforementioned drawbacks by limiting crop cultivation and 

production to a single growing season. The dehulling technique presents another challenge (Eke 

and Akobundu, 1993). 

 

2.10 Genetic Diversity within Species 

Breeders can create varieties for particular features, like improved biotic and abiotic resistance 

stressors and quality, more easily when there is genetic variety (Bhanu, 2017). Tannins are one 

type of anti-nutrient found in the seeds. Many methods, including (i) structural analysis, (ii) In the 

pregenomic era, biochemical characterization and assessment (allozyme), and (iii) In the 

postgenomic era, single nucleotide polymorphisms (SNPs) are especially useful for analyzing 

DNA (or molecular) markers, which are widely employed to measure genetic variation within and 

Between plant populations, saponins, hydrogen cyanide, phytic acid, and trypsin inhibitors. 

Markers may show comparable inheritance patterns to any other trait, such as codominant, 

recessive, or dominant. When a marker is codominant, it can differentiate between the hereditary 

patterns of homozygotes and heterozygotes. According to Govindaraj, Vetriventhan, and 

Srinivasan (2015), codominant markers typically provide more information than dominant 

markers. 



19  

2.11 Morphological Characterization of Sphenostylis stenocarpa 

Variation in plants occurs naturally. Man has been able to utilize this variation to enhance food 

production and breeding techniques. This variability can be between species or within species. Due 

to a variety of environments, plants have been able to evolve several physiological structures to 

survive their respective environments. Understanding their variation will aid our understanding of 

plants. To effectively use crop germplasm, plant materials with desirable features must be 

morphologically characterized (Santos, Pires & Correa, 2012). Morphological features have been 

extensively utilised in plants to distinguish species, families, and genera. The morphological 

characteristics that are most helpful for describing, characterizing, and evaluating farmed plants of 

economic significance have previously been chosen (Bidot Martínez et al., 2017). This can be 

achieved by visuals from the unaided eye. 

 

These characteristics include, among others, plant height, growth pattern, bearing habit, flowering 

pattern, disease resistance, inflorescence shape, color, kind of flowers, fruit shape, color, shoulder 

position, and seeds (polyembryonic or monoembryonic) (2010). The seeds of AYB reflect a lot of 

diversity in color, shape, and size, for instance, a crucial morphological factor in the classification 

of AYB is the color pattern of the seed. According to reports, seed-related characteristics have a 

significant role in defining the taxonomic and generic relationships of crops, including legumes 

(Adewale et al., 2012). During an examination of the African yam bean's physical characteristics 

and variability analysis (Aina et al., 2020). The Authors observed the significant mean square 

values for 19 quantitative agro-morphological variables assessed during two cropping seasons, 

they revealed notable variations in the performances of 50 AYB accessions and their respective 

significance. Eighty percent of the accessions had noticeable seed cavity ridges, which were 

primarily oval or rhomboid in shape. Two main groupings of African yam bean accessions were 

distinguished by their distinct eye color patterns. Four seed shapes were identified in the 

experiment; where five, 10% of accessions had round seeds, seventeen, 34% had oval seeds, 

nineteen, 38% had oblong seeds and nine, 18% had rhomboid seeds. In order to maximize AYB 

productivity, they also recommended that important factors be considered in breeding programs, 

such as the number of seeds per pod, number of pods per plant, weight of seeds per pod, and weight 

of seeds per plant. 



20  

In a different investigation on the physical description and identification of AYB germplasm from 

various ecological zones, Shitta et al. (2021). Several phenotypic characteristics showed 

considerable differences, indicating great variability throughout the germplasm under study. They 

discovered that growth habit was another qualitative characteristic that clearly distinguished the 

accessions; for instance, 56.8% of the plants had bushy habits, whereas 43.2% of the plants had 

erect patterns. Accessions with upright growth patterns were shown to yield more pods than bushy 

varieties. In contrast to the bushy type, the pods produced by erect types were arranged on branches 

so that they developed well above ground level, protecting them from damage by soil pests. 

 

The morphological diversity of African yam beans and their potential use in breeding and 

germplasm conservation were examined by Adewale et al. in 2012. The scientists observed that 

the presence or lack of variegation on the seeds clearly divided the AYB accessions into two 

groups, suggesting that seed color pattern is a crucial morphological factor in the AYB 

classification process. Additionally, they discovered a further set of 12 seed traits that are highly 

effective in differentiating AYB germplasm. In 2015, Ojuederie et al. evaluated forty African yam 

bean accessions. They characterized these accessions using 48 agro-morphological criteria during 

the course of two cropping seasons, from June to December in 2011 and 2012. Promising 

agronomic features in the accessions were found. For instance, TSs 52, TSs 51, TSs 68, TSs 26, 

19, TSs 66, AYB 9, and TSs 154 are examples of early flowering accessions. 



21  

 
 

 

Plate 2.1: Staked African yam bean plants carrying matured pods. 



22  

2.12 Molecular Characterization of Sphenostylis stenocarpa 

DNA sequence polymorphisms that occur naturally serve as the foundation for molecular markers. 

Plant genetic diversity can be assessed and characterized using molecular markers in a way that is 

independent of the environment in the fields of plant sciences such as Ecology, Evolution, 

Taxonomy, Phylogenics, and Genetics (Christensen et al., 2007). Commonly utilized genetic or 

DNA-based marker approaches include Simple Sequence Repeats (SSR), Amplified Fragment 

Length Polymorphism (AFLP), Random Amplified Polymorphic DNA (RAPD), and Restriction 

Fragment Length Polymorphism (RFLP). These methods are well-established, and it is known 

what their benefits and drawbacks are (Agarwal, Shrivastava & Padh, 2008). Molecular markers 

may or may not correlate with the phenotypic expression of a genetic characteristic, in contrast to 

morphological markers. Regardless of the growth, differentiation, development, or defense status 

of the cell, they are stable and detectable in all tissues, which gives them several advantages over 

traditional, phenotype-based alternatives. Furthermore, they are not confused by pleiotropic, 

environmental, or epistatic effects (Govindaraj, Vetriventhan & Srinivasan, 2015). 

 

DNA polymorphism can be quantified using a variety of molecular markers, which can be 

categorized into two groups: markers based on hybridization and markers based on Polymerase 

Chain Reaction (PCR) (Jonah et al., 2011). The development of genome-based discoveries and 

technological breakthroughs has improved molecular marker approaches. These sophisticated 

molecular marker approaches combine the best features of multiple fundamental approaches with 

additional methodology changes to improve resolution and sensitivity in identifying genetic 

discontinuity and distinctiveness (Agarwal, Shrivastava & Padh, 2008). Molecular tools have been 

notably used to unravel intra-specific diversity in AYB. For example, Adewale et al., in 2015, 

reported a study on a diversity analysis carried out on seventy-seven accessions of AYB utilizing 

AFLP markers. With the help of 5 primer combinations, 227 AFLP bands were generated. 

According to the study, the seed coat colors of TSs-98 (sun-burn brown) and TSs-104B (greyish 

white) differ from one another. Additionally, the existing strategy of classifying the 77 AYB 

accessions into four genetic groups based on DNA profiles and agro-morphological 

characterization supports parental selection and the formation of heterozygous breeding 

populations. 



23  

Shitta et al. (2016) examined the genetic diversity of African yam beans by examining the 

amplification capacity of thirty-six cowpea simple sequence repeat (SSR) primers across 67 

accessions' worth of AYB genomic DNA. The eight transferable SSR primers identified 55 

polymorphic fragments from the experiment. The cross-species transferability of SSRs and its 

application to comprehend diversity within the Fabaceae family were verified by the authors. They 

also reported some limitations to their study based on most of their accessions originating from 

Nigeria, which could only inform of limited diversity. They further recommended that more 

germplasm from different geographical regions be collected for a cleaner understanding of the 

AYB. A set of Nigerian accessions of African yam beans were examined for genetic variation 

using RAPD primers (Moyib et al., 2008). High genetic diversity was found using RAPD primers 

in a limited set of African yam bean accessions from Nigeria. While a minimum of six polymorphic 

primers was sufficient for genotype identification investigations, at least two RAPD primers may 

be utilized conveniently for genetic diversity studies in Nigerian accessions of AYB, but with low 

resolution. 

 

2.13 Concept of Yield Stability 

Yield instability is a significant problem, specifically used in African yam bean-producing areas 

characterized by multiple year variability in yield (Aremu et al., 2020). It has then resulted in the 

constant decline of AYB production or cultivation (Arogundade, Mu & Akinhanmi, 2016). The 

instability of yield can significantly affect breeding programs. The major aim of breeding programs 

is to attain stabilized germplasm and pinpoint breeding and testing conditions for the desired 

features (Adewale and Dumet 2011). Therefore, determining the extent of the G × Y interaction 

on yield stability in AYB is important because, for African farmers with limited resources, yield 

stability serves as a safeguard against crop failure. Therefore, identifying stable genotypes for seed 

production through extensive diversity across cultivars is crucial to the long-term viability of sub- 

Saharan Africa's smallholder agricultural system (Adewale and Dumet 2011). Field trials were 

carried out by Aremu et al. (2020) to investigate the performance of 23 AYB genotypes in four- 

year settings. After 5 plants from each row were picked independently, the seeds from every plant 

sampled in each plot were weighed and bulked, allowing for the calculation of the seed production 

per plant. To determine the significance of the genotypes, year, and genotype-by-year interaction, 

a combined analysis of variance (ANOVA) was carried out. 



24  

Before executing a combined ANOVA, the homogeneity of residual variances was assessed by the 

Bartlett test; genotype X Genotype X Environment interaction (GGE) biplot analyses were 

employed to ascertain the genotype stability across time in a quantitative and graphical manner. 

Another study conducted by Adewale et al., (2016) was conducted in multiple locations (Ikenne, 

Ibadan, Ubiaja and Mokwa) in Nigeria. They investigated the inheritance pattern and stability 

status of agronomic traits in African yam bean. It was discovered that there was no difference in 

days to seedling emergence (DSE) amongst the thirty genotypes under the combined analysis of 

variance. For the other attributes, there was variance among the 30 accessions that was extremely 

significant (P≤0.001). The 4 sites showed a significant difference (P≤0.001), while the four 

attributes showed an equally significant (P≤0.05) Genotype by Location (G x L) interaction. For 

every parameter, the G x L proportions were less than 10%; days to 50% flowering (D50F) and 

one hundred seed weight (100SW) had the lowest (1.78%) and greatest (5.17%) proportions, 

respectively. Stable genotypes with desired traits (such as earliness in DSE and D50F and high 

yield in 100SW and SWP) could be chosen as parents for additional enhancement of the trait of 

concern, as noted by Makinde & Ariyo (2012) and Yonas et al. (2014). The prospective 

performances and stability for the four features were not mutually exclusive, despite the 30 AYB 

accessions' varying stability for the four examined characters in the various conditions. 

 

2.14 Quantitative traits and Marker-Trait Association 

Quantitative traits are directed by multiple genes/loci (Zhang et al., 2020). Though these traits can 

be measured their phenotypic expressions are affected by the total activities of multiple genes and 

the interaction of these genes with a continuous distribution of phenotypes. A continuous 

distribution of phenotypes is produced by this interaction, which differs among individuals over a 

specified range (Sham et al., 2002). A QTL is a genetic locus whose alleles affect this variation. 

Since quantitative traits are often complex and influenced by several polymorphic genes and 

environmental factors, one or more QTLs may have an impact on a trait or phenotype. 

 

It is crucial to keep in mind that, in addition to genotype, environmental variables and gene- 

environment interactions can also influence phenotypic variation. (Members of the Complex Trait 

Consortium, 2003). Natural populations for linkage disequilibrium-based association mapping and 

experimental populations for linkage-based QTL mapping are the two most often utilised 



25  

techniques for QTL mapping (Olomitutu, 2023). Depending on the underlying principles, the 

various linkage-based QTL analysis techniques can be categorized into four main groups: (1) 

single-marker analysis in the absence of a linkage map; (2) interval mapping in the presence of a 

linkage map; (3) meta-QTL analysis; and (4) joint linkage and association mapping (Kulwal, 

2018). Bardol et al. (2013) combined linkage analysis (LDLA) with linkage disequilibrium data to 

map out QTL. In two sizable multi-parental datasets of maize (Zea mays L.), genotyped with 491 

markers and consisting of 895 and 928 testcross progenies made up of 7 and 21 biparental families, 

respectively, they were able to examine several methods for detecting QTL for four variables of 

agronomical value. 

 

Traditional linkage-based techniques were contrasted with two LDLA models that were based on 

the intensive genotyping of parental lines using 17,728 SNP. One model was based on a clustering 

strategy of parental line segments into ancestral alleles, while the other model was based on single 

marker information. Compared to traditional linkage models, which identified 49 and 44 QTL 

altogether, the two LDLA models frequently identified 60 and 52 QTL. The Association mapping 

method is used instead for the shortcomings of linkage mapping. It is based on linkage 

disequilibrium (LD) and was originally developed for human disease genes (Flint-Garcia et al., 

2005). Association mapping allows researchers to use different genomic platforms to exploit 

natural genetic diversity. At the moment, Genome-Wide Association (GWA) and Candidate Gene 

Association are the two association mapping approaches that were employed (Zhu et al., 2008). 

The correlation between polymorphisms in a population is referred to as LD. Because the 

genotypes of the genotyped markers and the functional variant have a strong correlation, the 

genotyped markers serve as proxies, or sentinels, for the functional variant (Myles et al., 2009). 

 

2.15 Gene and protein relationship 

A gene is an assembly of genomic sequences that together code for a logical group of functional 

products that may or may not overlap (Pesole, 2008). Genes can also be considered functional 

products in the genome that encode relevant information that influences genotypic and phenotypic 

characteristics. Every gene can exist in multiple forms known as alleles, and each gene is found 

on a chromosome. These alleles come from parents to offspring through sexual reproduction 

(Chen, 2020). 



26  

Table 1: Research gaps 

Title of Research Research 

Deficiency 

 

 

Gaps to be addressed References 

 
 

Bivariate analysis of the 

genetic variability among 

some accessions of African 

Yam Bean (Sphenostylis 

stenocarpa) 

Molecular analysis 

of the accessions 

was omitted 

Genotyping will be 

conducted for precise 

data confirmation 

Akinyosoye 

et al., 2017 

 

Genetic diversity in African 

yam bean 

accessions based on AFLP 

markers: towards a platform 

for germplasm 

improvement  and 

utilization 

Absence of 

phenotyping for 

environmental 

variation influence 

SNP markers will be 

used for better 

precision on diversity 

analysis with the help 

of DArTseq technology 

Adewale et 

al., 2015 

 

Genome-Wide Association 

Study Revealed SNP 

Alleles Associated with 

Seed Size Traits in African 

Yam Bean (Sphenostylis 

stenocarpa) 

Lack of reference 

genome sequence 

for quality 

alignment 

Marker assisted Traits 

association study for 

tuber forming 

accessions 

(Olomitutu et 

al., 2022) 

 

 

 
 



27  

CHAPTER THREE 

MATERIALS AND METHODS 

 
3.1 Research site 

Field study was conducted in Nigeria at the International Institute of Tropical Agriculture (Ibadan), 

for the 2023 and 2024 cropping season. 

 

3.2 Germplasm used 

One hundred AYB accessions in total, were acquired from the Genebank at IITA 

 

3.3 Experimental Design and Field Plot Management 

A 20 x 5 lattice pattern that was duplicated three times served as the experimental design. The date 

of planting was August 29, 2023. Single, 4-meter-long ridges placed 0.75 meters apart made up 

each plot. On ridges, seeds were spaced 0.5 meters apart and covered with Mancozeb 80% WP. 

Each hill had two seeds put in it and thinned to one plant per hill. After three weeks of sowing, 50 

kg P/ha of triple superphosphate fertilizer was applied, and the plants were staked. For the purpose 

of controlling floral and pod pests and fungal diseases, respectively, a fortnightly application of 1 

L/ha and 2 Kg/ha of Cypermethrin 30g/L+Dimethgoate 250 g/L EC and Mancozeb 80% WP was 

made from the beginning of blooming until harvest. In order to minimize weed interference after 

pre-emergence herbicide application, manual weeding was done on a regular basis. 



28  

3.4 Collection of Data 

Data were recorded using the AYB descriptor (Adewale and Dumet, 2011) as described below: 

1. Days to Flowering: Number of days from sowing to first plant stand begin to anthesize. 

2. Days to 50% flowering: Number of days from sowing to when 50% of the plant stand on 

the plot has flowered 

3. No. of pods per plant: The number of pods estimated using pods harvested from five 

representative plants. 

4. Total pod weight (g/plant): Total pod weight of each plant stand per accession. 

5. Pod Length: Estimate using the length of one randomly selected pod harvested from five 

representative plants. 

6. Pod width: Estimate using the width of one randomly selected pod harvested from five 

representative plants. 

7. No. of Seed per Pod: The number of seeds in a randomly selected pod harvested from five 

representative plants. 

8. Hundred seed count: Hundred count of randomly selected seeds taken from total seed 

yield/plant 

9. Total seed weight per plant: Weight of the total number of seeds taken from each plant 

10. Seed Thickness: Estimated using the veiner caliper 

11. No. of tubers per plant: Estimated using the number of tubers produced per plant. 

12. Total Tuber Weight: Estimated using the weight of tubers produced per plant. 

13. Tuber Length: Estimated using the length of the largest of the tubers harvested per plant. 

14. Tuber Width: Estimated using the width of the largest size of the tubers harvested per plant. 



29 

 

Table 3.1: AYB passport information displaying the nation of origin, the acquisition date, 

and the name of the collector. 

 

S/N Accession number Region/Origin 

in Nigeria 

Collection date Collector’s name 

1 TSs-481 North Central 20190131 NA 

2 TSs-629 South East 20190206 IITA 

3 TSs-609 South East 20190131 IITA 

4 TSs-597 South East 20190206 IITA 

5 TSs-441 North Central 20170621 NA 

6 TSs-91 North Central 19780718 Badra 

7 TSs-567 South South 20190204 IITA 

8 TSs-598 South East 20190206 IITA 

9 TSs-471 North Central 20190204 NA 

10 TSs-138 North Central 19761201 NA 

11 TSs-445 North Central 20170621 NA 

12 TSs-268 North Central 19761201 NA 

13 TSs-133 North Central 19761201 NA 

14 TSs-56 North Central 19730403 L.Igbokwe 

15 TSs-63 North Central 19730514 Okigbo 

16 TSs-201 North Central 19761201 NA 

17 TSs-121 North Central 19791201 Dr. N. Q. Ng 

18 TSs-558 South East 20190202 IITA 

19 TSs-642 South East 20190206 IITA 

20 TSs-556 South East 20190202 IITA 

21 TSs-550 South East 20190202 IITA 

22 TSs-560 North 20190202 IITA 

23 TSs-95 South East 19780718 Badra 

24 TSs-427 North Central 20170621 NA 



30 

 

 

25 TSs-157 North Central 19761201 Machuka 

26 TSs-647 South East 20190207 IITA 

27 TSs-431 North Central 20170621 NA 

28 TSs-24 North Central 19721123 Kanti Rawal 

29 TSs-421 North Central 19761201 NA 

30 TSs-511 South East 20190131 IITA 

31 Tss-366 North Central 19761201 NA 

32 TSs-547 South East 20190202 IITA 

33 TSs-53 North Central 19730404 D. Ajoku 

34 TSs-153 North Central 19761201 Dr. J. Machuka 

35 TSs-562 South East 20190204 IITA 

36 Tss-26 North Central 19721123 Kanti Rawal 

37 TSs-564 South South 20190204 IITA 

38 TSs-537 South East 20190201 IITA 

39 TSs-437 North Central 20170621 NA 

40 TSs-581 South South 20190205 IITA 

41 TSs-47 North Central 19730112 R.J. Williams 

42 TSs-42 North Central 19721204 Rawal 

43 TSs-275 North Central 19761201 NA 

44 TSs-592 North Central 20190206 IITA 

45 TSs-527 South East 20190131 IITA 

46 TSs-463 North Central 20190204 NA 

47 TSs-554 South East 20190202 IITA 

48 TSs-9 North Central 19720801 C.N. Aniagu 

49 TSs-548 South East 20190202 IITA 

50 TSs-571 North Central 20190204 IITA 

51 TSs-142 North Central 19761201 Dr. J. Machuka 

52 TSs-101 North Central 19791201 NA 

53 TSs-588 South South 20190205 IITA 

54 TSs-61 North Central 19730514 Okigbo 



31 

 

 

55 TSs-84 North Central 19780718 Badra 

56 TSs-601 South East 20190206 IITA 

57 TSs-618 South South 20190205 IITA 

58 TSs-100 North Central 19791201 NA 

59 TSs-432 North Central 20170621 NA 

60 TSs-525 South East 20190131 IITA 

61 TSs-551 South East 20190202 IITA 

62 TSs-593 South East 20190206 IITA 

63 TSs-249 North Central 19761201 NA 

64 TSs-517 South East 20190131 IITA 

65 TSs-150 North Central 19761201 Dr. J. Machuka 

66 TSs-166 North Central 19761201 NA 

67 TSs-627 South East 20190206 IITA 

68 TSs-651 South East 20190207 IITA 

69 TSs-424 North Central 19761201 NA 

70 TSs-508 South East 20190131 IITA 

71 TSs-209 North Central 19761201 NA 

72 TSs-14 North Central 19721120 Kanti Rawal 

73 TSs-197 North Central 19761201 NA 

74 TSs-276 North Central 19761201 NA 

75 TSs-161 North Central 19761201 NA 

76 TSs-465 North Central 20190204 NA 

77 TSs-117 North Central 19791201 Dr. N. Q. Ng 

78 TSs-553 South East 20190202 IITA 

79 TSs-536 South East 20190201 IITA 

80 TSs-482 North Central 20190131 NA 

81 TSs-40 North Central 19721125 Kanti Rawal 

82 TSs-293 North Central 19761201 NA 

83 TSs-371 North Central 19761201 NA 

84 TSs-631 South East 20190206 IITA 

85 TSs-522 South East 20190131 IITA 



32  

 

86 TSs-320 North Central 19761201 NA 

87 TSs-4 North Central 19720801 C.N. Aniagu 

88 TSs-430 North Central 20170621 NA 

89 TSs-458 North Central 20190206 NA 

90 TSs-111 North Central 19791202 Dr. N. Q. Ng 

91 60B North Central NA NA 

92 TSS-44C North Central NA NA 

93 TSs-505 South East NA NA 

94 TSs-502 South East NA NA 

95 59B North Central NA NA 

96 TSs-340 North Central NA NA 

97 TSs-673 South East NA NA 

98 TSs-670 South East NA NA 

99 TSs-673 South East NA NA 

100 TSs-680 South East NA NA 



33  

3.5 Equipment 

The equipment used in this research were lyophiliser (Fisher Scientific, USA); Vortex(Model 

F202A0175 Scientifica, Italy); Water bath (Model 1102, Gesellschaft fur Labortechnik mbH, 

Germany); Centrifuge (Model 5418, Eppendorf, Germany); Genogrinder (Fisher Scientific, USA); 

Test tube, Eppendorf, Germany). 

 

3.6 Reagents 

The reagents used in this research were of analytical grade and they included betamercaptoethanol, 

cetyl trimethyl ammonium bromide (CTAB) extraction buffer, 70% ethanol, isoamylalcohol, 

chloroform, authoclaved distilled water, ice cold isopropanol; agarose tablets, RNAse A, 

Ethylenediamine Tetraacetic acid (TE) buffer ethidium bromide, Mancozeb, phenol-sulfuric acid. 

 

3.7 Methods 

 

3.7.1 Phenotypic Analysis 

A total of one hundred accessions were regenerated on the field at the International Institute of 

Tropical Agriculture, Ibadan, Oyo State, Nigeria. The planting was done for one cropping seasons 

i.e., 2023/2024 (August to February). The field plots comprised single 4 m long ridges spaced 0.75 

m apart. A total of five plants per accession were used for the collection of data. 

The young leaves were harvested at three weeks old for genotyping while the emerging plants 

were staked and twinned after three weeks of planting. 

 

3.7.2 Genotyping 

Three to four young leaves of three weeks old plants were collected per accession for genomic 

DNA extraction utilizing the updated Cetyltrimetyl Ammonium Bromide (CTAB) method (Doyle 

and Doyle, 1990). The fresh young leaves were collected into well-labelled paper bags and placed 

on ice for transportation to the laboratory. All the samples were stored in the freezer for 24 hours 

after which they were freeze-dried for 72 hours using a lyophiliser (Fisher Scientific, USA). Each 

accession (100 mg) was put into 2 ml Eppendorf tubes which contained steel balls for grinding. 

The samples were ground using a genogrinder (Fisher Scientific, USA). After grinding, the steel 

balls were removed and 600 micro-litres CTAB extraction buffer was put into each of the one 

hundred tubes for the DNA extraction. After that, the samples were submerged in water at 65 

degrees Celsius for 30 minutes with tender rocking and alternating inversions every 10 minutes. 



34  

After 30 minutes, the tubes containing the samples were removed from the water bath and allowed 

to cool for 5 minutes in the fume hood. After cooling 600 micro-litres of chloroform: isoamyl 

alcohol (24:1) was added into each of the samples and mixed continuously by shaking. The 

samples were then centrifuged at 3500 rpm for 10 minutes. The supernatants were transferred 

gently into freshly labelled tubes. The chloroform: isoamyl alcohol wash was carried out twice. 

The supernatant for each sample (600 micro-litres) was transferred into freshly labelled tubes and 

400 micro-litres of ice-cold isopropanol was added to each sample. The samples were mixed gently 

and kept in the -80 degree Celsius freezer for 90 minutes. The samples were removed from the 

freezer for the samples to thaw and then the samples were then centrifuged at 3500 rpm for 30 

minutes. Following centrifugation, the supernatants were then decanted and 400 micro-litres of 

70% ethanol was added to each tube for easy washing of the pellets by flapping the tubes gently. 

The samples were then centrifuged at 3500 rpm for 10 minutes. The ethanol wash was repeated 

twice. The supernatant for each sample was decanted and pellets were air-dried in the fume hood 

for complete evaporation of the ethanol. Each air-dried DNA sample was suspended with 95 

micro-litres low salt Tris-EDTA (TE) buffer and 5 micro-litres of RNAse A. The samples were 

incubated at 37 degrees Celsius for 45 minutes. 

3.7.2.1 Quality Control 

 

Agarose gel electrophoresis was used to determine the quality of genomic DNA. Genomic DNA 

(2 micro-litres) mixed with 2 micro-litres ethidium bromide was loaded on 1% agarose gel and ran 

at 100 V for 30 min. One micro-litre of genomic DNA was measured using a nano- 

spectrophotometer to ascertain the DNA concentration. 

3.7.2.2 DArT-Seq 

High-throughput genotyping was carried out at DArT Private Limited, University of Canberra, 

Monana St., Bruce, ACT 2617, Australia (www. diversityarrays.com) using the Diversity Array 

Technology sequence (DArTSeq). The 100 AYB accessions were genotyped to yield molecular 

markers for seed and tuber traits. 

http://www/


35  

3.7.2.3 SNP Filtering 

Reads and tags identified in the individual African yam bean sequences were aligned to the 

Common bean (Phaseolus vulgaris) genome. A total of 5142 SNP markers were obtained from 

DArTSeq and filtered based on the following criteria: monomorphic markers, 70% call rate for 

markers, 95% reproducibility, 25% missing data, and 1% minor allele frequency (MAF). After 

filtering, 2254 SNP markers were used for GWAS analysis. 

3.7.2.4 Statistical Analysis 

The values of the best linear and unbiased estimators (BLUEs) were estimated across two years 

using the META-R software. The values of the BLUE estimates across both years were employed 

for the Genome-Wide Association Study (GWAS) analysis using the mixed linear model to reduce 

false positives as well as type I and II errors (Adewale et al., 2020). Also, Manhattan plots were 

displayed using the GAPIT package. A GWAS threshold of p-values were used for Q_Q plot (Li 

et al., 2019; Ding et al., 2022). 

3.7.2.5 Principal Component Analysis (PCA) and Cluster Analysis 

PCA was done to find out which features were most useful in differentiating between accessions. 

Eigenvalues of principal components (PCs) greater than 1.0 were kept in the analysis. To provide 

more light on the connections between the PCs and the variables, For the first two PCs, a PCA 

biplot analysis was carried out. Using Ward minimal variance method, cluster analysis (combining 

pairwise Mahalonobis genetic distances and constellation plot) across clusters) was carried out 

using c (Ward, 1963). Both PCA and cluster analysis were carried out using the R statistical 

software. 

3.7.2.6 Inter-trait Relationship 

To determine the inherent relationship between paired traits, phenotypic (𝒓𝚙) genotypic (𝒓g) 

correlation coefficients were estimated using the R statistical tool. version 4.3.2. For further 

information on the interrelationships among the traits studied, path coefficient analysis was done 

to determine the direct and indirect effects of correlation coefficients were used in the path analysis 

as suggested by Kang et al. (1991) to avoid spurious association in the phenotype as a result of 

artificially created relationships among traits in a pathway. 



36  

Predicted on the p-value distribution for all the features, trait-marker relationships were considered 

significant at P-values of -log (p) = 4. (Mogga et al., 2018; Adewale et al., 2020). Quantile-Quantile 

plots and Manhattan were constructed accordingly. All analyses were performed in Tassel 

software. Molecular markers are known as reference landmarks for genes in the genome. To further 

validate the significant marker-trait association found in this study using related legume genomes. 

The integrated location analysis (Phaseolus vulgaris G19833 v2.0) on the genome database in the 

Legume information system for common beans (Phaseolus vulgaris), a blast search was conducted 

for the trimmed nucleotide sequence of important AYB markers (Dash et al., 2016). The search 

was also extended to syntenies of allied legumes (Glycine max 2.0, Vigna angular 3.0, and Cajanus 

cajan 1.0). The scroll was magnified to 1 Mb (500 Kbp up and downstream) to ascertain whether 

the nearby genes and the enzymes they encode affect the properties of interest. This was done due 

to lack of AYB reference genome limits candidate gene mapping. 



37  

CHAPTER FOUR 

RESULTS 

 

 

4.1 Phenotype Analysis 

 

4.1.1 : Seed 

The results of the study showed a consistency of seed production in majority of the accessions 

across the replicates. TSs-571 showed the highest yield per plot. The clustering pattern of the seed 

yield was indicated by four clusters below the intercept line. With cluster III having accessions 

with the highest seed yield. 

Multiple traits such as Number of Pods, Hundred Seed Count, Total Seed Weight per Plant, and 

Total Pod Weight indicate a close association with each other showing a strong indication with the 

accessions and yield. Total Pod Weight and Total Seed Weight per Plant had the highest level of 

correlation standing at 0.98, while Number of Pods and Pod Width had the lowest level of 

correlation. 



38  

 

 

 

 

 

Table 4.1: Shows the Seed raw data of the accessions and their replicates 

Accession    REP     Block              NoPods         TPodwgth     PodLngth        Podwdth       NoSdPp       Hund_Sdcount    TSwghtPPl    SdThck 

59B 1 4 5.6 29.38 24.58 8.936 14.4 62 18.56 6.992 

59B 2 9 5.7 37.85 25.1 9.33 16.5 74.75 20.925 7.4025 

59B 3 12         

60B 1 1 5 12.3 21.525 6.33 10 44 7.05 5.38 

60B 2 8 6.4 23.7 23.18 3.424 16.8 69.2 15.4 7.164 

60B 3 11         

TSs-100 1 4 1 1.4 19.5 8.12 6 6 0.1 2.99 

TSs-100 2 8         

TSs-100 3 13 3.5 17.35 22.75 10.155 12.5 39 9.85 7.255 

TSs-101 1 3 7.75 33.675 23.375 8.685 11.5  21.275 6.305 

TSs-101 2 7 4 23.375 26.25 9.1375 17 54.25 14.7 6.6575 

TSs-101 3 14         

TSs-111 1 5 1 2.8 24.1 9.89     

TSs-111 2 8 3.5 11.75 19.2 7.65 9.5 28.5 5.3 6.8 

TSs-111 3 14 5 29.7 27.8 8.43 15 69 19.4 6.95 

TSs-117 1 5 3.5 16.725 22.175 8.725 14.75 47.25 10.3 6.0025 

TSs-117 2 8 5 23.36 24.38 8.866 14.6 64.6 14.62 6.8 

TSs-117 3 11 1.5 7.65 24.35 9.62 16.5 16 4.35 5.915 

TSs-119A 1 3 3 12.18 22.94 8.896 11  9.8 7.184 

TSs-119A 2 7 6.8 28.32 23.92 9.108 12.6 52.6 16.62 7.068 

TSs-119A 3 14 6.6 27.76 25.76 8.664 13.4 50.06 17.76 7.288 

TSs-121 1 1 1 3.3 19.5 8.92 6 6 2.3 7.4 

TSs-121 2 6 4.7 24.7 21.3 9.01 10.7 47.7 14.8 7 

TSs-121 3 14 1 7 23.3 8.93 16 16 5.3 7.36 

       

 



39  

  

 
 
 
 
 
 

 
TSs-197 1 4 2 11.6 26.7 8.28 16 26 7.5 6.73 

TSs-197 2 9 9.2 53.58 26.78 9.752 17.4 83.4 35.2 7.052 

TSs-197 3 13 4.8 20.5 23.64 9.452 13.2 47.8 12.12 7.022 

TSs-201 1 1 3 14.95 24.9 8.55 13 46 10.3 5.395 

TSs-201 2 10 7.5 52.925 28.5 10.04 17.5 86 32.7 6.6275 

TSs-201 3 11 2.5 12.175 22.475 9.2675 12.5 32.25 8.366667 6.555 

TSs-209 1 4 2.5 12.95 24 8.59 13 18.5 4.1 6.26 

TSs-209 2 6 5.666667 26.06667 25.9 9.42 13.66667 53 15.9 6.836667 

TSs-209 3 12 5.8 19.52 23.64 8.736 13.4 38 12.4 6.812 

TSs-24 1 2 4.2 19.06 22.64 9.078 12 47.6 11.46 6.206 

TSs-24 2 10         

TSs-24 3 14 4.8 32 23.66 9.492 13.8 57 18.7 6.15 

TSs-249 1 4 2 7.5 22.2 8.1 11  4.8 6.14 

TSs-249 2 9 6.5 35.6 27.075 8.6375 11.25 68 23.8 6.5325 

TSs-249 3 12 4.25 20.8 24.8 8.7525 14.75 49.25 13.525 6.59 

Tss-26 1 2 8 40.3 25.4 8.9 16 100 26.2 6.17 

Tss-26 2 10 6 27.46667 22.9 9.223333 12.66667 68.66667 22.56667 7.166667 

Tss-26 3 12         

TSs-268 1 1 7 27.2 23.76 7.746 11.8 63 17.18 6.056 

TSs-268 2 9         

TSs-268 3 15 14 52.1 22.6 8.108 13.4 88.8 32.08 6.55 

TSs-275 1 3 2 10.03333 20.93333 9.413333 11 23 5.8 6.793333 

TSs-275 2 8 4.2 17.92 24.34 8.844 13 39.8 10.7 6.054 

TSs-275 3 14 3.333333 11 23.9 9.193333 10.66667 29 5.9 6.683333 

           

           

           

           

           

           

           

 



40  

 

TSs-320 3 15 5.333333 22.4 23.23333 9.926667 12.66667 59 13.6 6.423333 

Tss-340 1 5 5.6 27.42 22.78 9.178 13.2 72.2 17.88 7 

Tss-340 2 7 3.8 16.9 20.88 6.244 11.2 40.2 10.24 6.758 

Tss-340 3 12         

Tss-366 1 2 3.5 10.875 23.075 8.0925 12 42.5 6.875 5.7075 

Tss-366 2 7         

Tss-366 3 11         

TSs-371 1 5 7 36.88 24.82 9.158 14.6 71 23.38 6.706 

TSs-371 2 10         

TSs-371 3 12 5.75 26.525 26.05 8.9 14.5 61.25 16.75 6.975 

TSs-4 1 5         

TSs-4 2 9 8.6 38.28 23.36 8.366 14.6 82.8 23.66 6.802 

TSs-4 3 14 4.4 25.52 24.46 8.482 16.2 61.8 17.6 6.888 

TSs-40 1 5 4 32 24 9.07 16 56 22.1 6.93 

TSs-40 2 10 3 11.46667 24.03333 8.683333 13 26.33333 6.533333 7.016667 

TSs-40 3 12 2.666667 13.03333 25.83333 8.436667 15.33333 32.33333 8.5 6.383333 

TSs-42 1 3 6.55 32.575 25.075 8.18 13.25 50.25 17.95 6.1725 

TSs-42 2 7 8.333333 42.43333 25.1 9.386667 15.33333 91 27.6 7.316667 

TSs-42 3 11 8 40.85 25.5 9.435 17 94.5 26.45 6.865 

TSs-421 1 2 1.5 8.8 16.35 9.765 10.5 14.5 4.85 6.655 

TSs-421 2 6 2.75 12.25 24.05 10.235 10.5 19.75 6.075 6.945 

TSs-421 3 11 2.666667 11.86667 24.13333 9.76 12.33333 20.66667 6.233333 6.96 

TSs-424 1 4 9.2 46.56 24.36 9.59 13.8 70.4 30.34 7.2 

TSs-424 2 8 1.5 8.45 23.65 9.56 14 18.75 5.05 7.37 

TSs-424 3 11 8.75 49.025 24.65 9.015 14 82 31.8 6.9925 

TSs-427 1 2 3.666667 15.7 22.26667 9.5 13.33333 52 12.76667 6.14 

TSs-427 2 10         

TSs-427 3 14 5.666667 26.03333 18.73333 8.226667 14 70 19.9 6.783333 

TSs-430 1 5 5 19.83333 22.73333 8.846667 15 54 13.13333 7.186667 

TSs-430 2 6 3.5 16.65 20 8.595 12.5 35 9.35 7.49 

           

           

           

 



41  

 

TSs-445 2 6 5.25 18.5 20.95 8.64 11.75 46 10.45 6.64 

TSs-445 3 13 5 25.3 23.02 9.17 12.4 49.6 16.62 6.902 

TSS-44C 1 1 2.4 10.84 20.62 9.006 11.6 27.2 6.62 6.62 

TSS-44C 2 10         

TSS-44C 3 11 4.8 21.2 24.42 9.068 13 45.8 12.98 6.834 

TSs-458 1 5 2 4.4 15.9 7.65 10 10 2.6 6.08 

TSs-458 2 6 3.2 17.44 25.44 9.084 14.2 36.8 11 6.506 

TSs-458 3 14 3 15.4 24.8 8.4425 14.5 42.5 10.225 5.805 

TSs-463 1 3 4.333333 23.96667 26.2 9.106667 16.33333 61.33333 16.93333 6.446667 

TSs-463 2 9         

TSs-463 3 15 8.4 41.48 26.98 9.1 16.8 82.8 30.24 6.612 

TSs-465 1 5 2 6.9 21.4 8.75 5 18 3.1 6.41 

TSs-465 2 10 9.2 34.44 21.86 8.994 12.6 84.2 21.5 6.656 

TSs-465 3 14 4.5 14.325 22.85 7.83 14.25 51.25 9.475 6.485 

TSs-47 1 3 1 4.3 24.6 8.985 11.5 11.5 2.55 6.37 

TSs-47 2 7 5.75 36.175 24.85 8.9625 14.75 71.5 22.15 6.205 

TSs-47 3 14 4.666667 26.66667 24.73333 7.843333 17.33333 70.66667 18.26667 6.396667 

TSs-471 1 1 6.325 15.575 25.45 9.79 12 29.25 9.375 6.36 

TSs-471 2 7 1.5 8.15 25.05 9.84 13.5 20 4.6 6.695 

TSs-471 3 13         

TSs-481 1 1 1 1.95 13.3 9.215 6 6 1.15 6.17 

TSs-481 2 9 4.5 18.85 21.35 9.82 12 43 11.75 7.405 

TSs-481 3 13 2.5 8.25 16.75 9.15 7.5 20 4.2 6.365 

TSs-482 1 5 3.8 18.58 22.24 10.3225 10 34.8 10.68 7.364 

TSs-482 2 6 6 24.84 23.2 9.64 11 55.4 14.74 7.084 

TSs-482 3 12 4.666667 32.66667 22.7 10.38333 12.33333 55.33333 20.03333 7.26 

TSs-502 1 4 4.8 17.12 20.3 9.376 11.2 39 10.62 7.362 

TSs-502 2 6 3.8 13.02 21.44 9.398 9.2 25.8 7.14 6.532 

TSs-502 3 12 14.5 63.65 23.55 9.2025 13.25 61.25 36.2 6.985 

           

           

           

           

           

           



42  

 

TSs-525 1 4 2 16.875 27.75 8.58 14.25 28.75 9.65 7.29 

TSs-525 2 8 4 21.94 25.74 9.336 13.4 35 10.66 6.762 

TSs-525 3 14 3.4 27.22 27.86 10.262 13.8 44 18.3 7.168 

TSs-527 1 3         

TSs-527 2 9 2.5 17.55 28.05 10.235 18.5 33.5 10.9 5.75 

TSs-527 3 11         

TSs-53 1 2 6 28.025 22.125 7.9825 11.75 66.25 17.925 6.35 

TSs-53 2 7 4.4 16.92 21.56 8.826 11.2 35.6 9.38 5.86 

TSs-53 3 15 11.2 57.22 26.62 8.564 15 100 41.78 6.926 

TSs-536 1 5 3 10 26.5 8.57 17 33.5 6.8 6.43 

TSs-536 2 8         

TSs-536 3 13 7.333333 41.5 24.3 9.203333 12.33333 73.33333 25.86667 6.646667 

TSs-537 1 2         

TSs-537 2 7 3 9.8 23.7 7.61 9 24 5.9 6.11 

TSs-537 3 13 4 29.3 25.725 9.93 13.5 41.25 18.325 6.8325 

TSs-547 1 2 3.5 16.8 22.35 11.085 9.5 30.5 9.65 7.345 

TSs-547 2 8 8.6 49.3 25.96 9.566 14.2 69.6 31.56 7.524 

TSs-547 3 14 4 25.8 29.83333 11.2 13.33333 47.66667 14.93333 7.13 

TSs-548 1 3 2.333333 10.26667 21.86667 9.2 8.666667  4.633333 6.62 

TSs-548 2 9         

TSs-548 3 11 3 25 29.85 11.22 18.5 47 14.95 7.86 

TSs-550 1 2 2.333333 10.56667 25 10.2 8.666667 15 5.3 7.253333 

TSs-550 2 9 5 31.5 26.75 11.255 11.75 52 18 7.9 

TSs-550 3 15 3.25 21.7 25.775 11.5225 12.75 35.75 11.85 7.53 

TSs-551 1 4 2.2 14.06 27.24 10.632 11.8  7.82 7.84 

TSs-551 2 8         

TSs-551 3 11 4 30.875 30.65 10.99 13.25 50 17.15 7.8525 

           

           

           

           

           

           

           

           

           



43  

 
 
 
 
 
 
 
 
 
 TSs-560 3 13 3 11.9 22.2 8.62 16 24 5.8 6.23 

TSs-562 1 2 2.5 8.45 19.95 9.29 7 15.5 3.5 6.045 

TSs-562 2 6 7 45.52 25.3 10.094 16.4 84.2 28.86 7.726 

TSs-562 3 13 2.666667 20 28.06667 9.393333 11.33333 36.66667 11.6 7.967767 

TSs-564 1 2         

TSs-564 2 10         

TSs-564 3 13         

TSs-567 1 1 1 0.6 13.8 9.42 0 0 0 0 

TSs-567 2 10         

TSs-567 3 15 1.333333 8.566667 23.86667 9.38 13 18 5.533333 6.893333 

TSS-56A 1 5 1 4.5 21.2 8.26 9  2.3 6.46 

TSS-56A 2 9         

TSS-56A 3 11         

TSs-571 1 3 4.2 16.46 19.68 8.35 12.2  10.68 6.808 

TSs-571 2 9         

TSs-571 3 13 9 49.9 23.9 9.14 14.5 68 34.85 7.12 

TSs-581 1 3 2 8.6 20.8 8.56 18 29 5.3 6.54 

TSs-581 2 7 4.666667 15.23333 20.93333 9.29 12.66667 29.66667 7.433333 6.666667 

TSs-581 3 15         

TSs-588 1 3 3.5 13.7 23.25 8.675 14.5 38.5 8.7 5.87 

TSs-588 2 10 2 6.5 21.2 7.71 14 28 4.3 5.24 

TSs-588 3 12         

TSs-592 1 3 6.2 27.26 21.98 8.72 14.8 75.2 17.4 6.212 

TSs-592 2 10 2.25 9.025 23.75 8.4625 13.5 24 5.525 6.5325 

TSs-592 3 12         

TSs-593 1 4 1 2.3 15.65 7.44 7 7 2.3 5.645 

TSs-593 2 9 1.666667 9.3 17.66667 9.726667 10.33333 20.66667 5.666667 7.53 

TSs-593 3 11 4.5 25.2 27.35 9.435 16 61.5 15.7 8.05 

           

           

           

           

           

           

           

           

           



44  

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

TSs-61 2 6 2 5.4 26.1 7.62 7 14 2.4 4.88 

TSs-61 3 11 4.75 31.9 26.525 9.3 15 64.75 19.85 6.9325 

TSs-618 1 4         

TSs-618 2 6 13.6 5.2 22.5 9.685 14.5 14.5 3.15 5.65 

TSs-618 3 12 2 9.8 21.9 9.28 12 21 5.6 6.4 

TSs-627 1 4 3 14 23.2 9.56 9  9.8 7.32 

TSs-627 2 6 3 16 28 9.53 17 33 9.5 7.07 

TSs-627 3 14         

TSs-629 1 1 4 13.2 19.66667 8.85 9 34.33333 8.933333 6.456667 

TSs-629 2 6 4.333333 23.06667 22.93333 9.266667 15.33333 46 14.1 7.126667 

TSs-629 3 13 13 58.36 23.96 9.938 15.4 81.4 36.54 6.814 

TSs-63 1 1 3.6 14.52 23.78 8.184 15.6 45 10.1 5.554 

TSs-63 2 9 4.5 26.8 23 8.415 18.5 54 17.1 7.43 

TSs-63 3 12 6.4 38.96 24.58 9.754 17.8 61.6 24.64 7.132 

TSs-631 1 5 1 4.7 21.6 8.34 8 8 2.8 6.98 

TSs-631 2 7         

TSs-631 3 12 6 43.2 29.2 9.8 11 72 23.5 7.19 

TSs-642 1 2 2 4.2 25.1 8.67 10 18 1.8 5.03 

TSs-642 2 9         

TSs-642 3 13 5.5 38.15 32.6 10.34 9.5 60.5 19.15 6.76 

TSs-647 1 2 2.25 6.35 19.275 9.0825 14.75 22.25 3.9 5.9275 

TSs-647 2 7         

TSs-647 3 11 2 6.2 17.5 9.01 11 21 3.9 5.85 

TSs-651 1 4 4 19 24.2 7.7 18 56 12.5 6.6 

TSs-651 2 7 3 16.6 23.5 9.38 15 37 10.9 7.71 

TSs-651 3 15 3 8.25 23.25 8.145 12.5 35 5.45 6.525 

TSs-84 1 3 3.2 13.66 22.2 9.318 13.4 39 8.68 6.574 

TSs-84 2 7 6.6 29.42 23.16 9.118 12.8 52.8 17.24 6.718 

TSs-84 3 12 3.6 19.84 24.84 9.024 15.8 45.4 12.34 6.312 

           

           

           

TSs-91 1 1 2 1.1 13.1 5.74 6 11 0.6 3.9 

TSs-91 2 9         

TSs-91 3 14 10 65.6 30.3 7.51 21 100 44.6 7.6 

TSs-95 1 2 3 15.7 25.4 9.57 14.5 40 10.2 6.86 



45  

 
 
 
 
 

 
NoPods   No of Pods 

                         Tpodwgth   Total Pod weight 

                         PodLngth   Pod Length 

                         Podwdth   Pod width 

                         NoSdPpd   Number of Seed per Pod  

                         Hund_Sdcount    Hundred Seed Count 

 

           TSdwghtPPl  Total Seed Weight Per Plant 

 

            SdThck   Seed Thickness 



46  

 

 

 

 

 

 

 

 

 

Figure 4.1: The cladogram showing the clustering pattern of the accessions based on seed yield. 

III 



47  

 

 

 

 

 

 

 

 

 

 

Figure 4.2: Biplot analysis of seed yield performance and their influencing traits 



48  

 

 

 

 

 

 

 
 

 

Figure 4.3: Identification of correlation of traits on the seed yield 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 



49  

 

 

 

4.1.2 : Tuber 

The results of the study showed an inconsistency of tuber production in majority of the accessions 

across the replicates. TSs-421 and TSs-209 showed the highest yield per plot. The clustering 

pattern of the tuber yield was indicated by five clusters below the intercept line. With cluster IV 

having accessions with the highest tuber yield. Multiple traits such as Blue_Tuber_weight_group, 

Blue_Number_Tuber_group indicate a close association with each other showing a strong 

indication with the accessions and yield. Blue_Tuber_weight_group and 

Blue_Number_Tuber_group had the highest level of correlation standing at 0.62, while Blue_ 

Tuber_Length_group and Blue_Number_Tuber_group had the lowest level of correlation at -0.05. 

There was no correlation between tuber traits and seed traits. Results were grouped into categories 

based on their result performance across the reps. 



50  

Table 4.2: Shows the raw tuber data transformed into categorized data 

S/N Accession Category_P_A Category_Amount Category_Wgt Category_lngt Category_Wdt 

1 59B 2 2 2 3 3 

2 60B 2 2 2 2 3 

3 TSs-100 1 1 1 1 1 

4 TSs-101 2 2 2 3 3 

5 TSs-111 2 2 2 3 4 

6 TSs-117 3 2 2 3 4 

7 TSs-119A 2 2 2 2 2 

8 TSs-121 3 3 4 4 4 

9 TSs-133 3 2 1 3 4 

10 TSs-138 3 2 4 3 5 

11 TSs-14 1 1 1 1 1 

12 TSs-142 1 1 1 1 1 

13 TSs-150 2 2 2 3 3 

14 TSs-153 2 2 2 3 3 

15 TSs-156A 1 1 1 1 1 

16 TSs-157 1 1 1 1 1 

17 TSs-161 4 4 4 4 4 

18 TSs-166 2 2 2 2 2 

19 TSs-197 4 2 2 3 4 

20 TSs-201 1 1 1 1 1 

21 TSs-209 2 2 2 3 3 

22 TSs-24 4 2 3 3 5 

23 TSs-249 4 4 9 4 4 

24 TSs-26 3 3 2 2 3 

25 TSs-268 2 2 3 3 4 

26 TSs-275 1 1 1 1 1 

27 TSs-276 2 2 2 3 3 

28 TSs-293 2 2 2 3 3 

29 TSs-320 1 1 1 1 1 

30 TSs-340 2 2 2 4 2 

31 TSs-366 3 3 3 3 4 

32 TSs-371 2 2 2 3 3 

33 TSs-4 1 1 1 1 1 

34 TSs-40 4 3 3 3 4 

35 TSs-42 4 5 8 3 5 

36 TSs-421 2 2 2 2 2 

37 TSs-424 3 2 2 3 4 

38 TSs-427 1 1 1 1 1 

39 TSs-430 1 1 1 1 1 

40 TSs-431 3 2 2 2 3 

41 TSs-432 2 2 1 2 3 

42 TSs-437 3 3 6 3 5 



51  

 

43 TSs-441 1 1 1 1 1 

44 TSs-445 2 2 2 4 3 

45 TSS-44C 4 3 2 3 3 

46 TSs-458 1 1 1 1 1 

47 TSs-463 2 2 1 3 3 

48 TSs-465 2 2 2 3 3 

49 TSs-47 4 2 2 3 4 

50 TSs-471 4 2 3 4 5 

51 TSs-482 2 2 3 3 3 

52 TSs-502 1 1 1 1 1 

53 TSs-505 1 1 1 1 1 

54 TSs-508 2 2 3 3 5 

55 TSs-511 3 2 2 4 4 

56 TSs-517 2 2 2 3 3 

57 TSs-522 1 1 1 1 1 

58 TSs-525 1 1 1 1 1 

59 TSs-527 4 3 6 4 6 

60 TSs-5