Annual Report ProJect SB-2 2004 r~r~Jif U'll A p 1 ff >CtON Y ¡,._ uMLfil AI..IUfi 1) Summary Conservat10n and Use of Tropical Genetlc Resources N ovember, 2004 ProJect SB-2 Content Conservatton and Use ofTroptcal Genettc Resources 2 A PROJECT OVERVIEW 2 B Pnnc1pal Collaborators 3 C Budget 6 D CIAT SB 2 ProJect Log Frame (2005 2007) 7 E Current SB 2 Investigators DISCiplme pos¡tJon and time fract10n 18 F H1gh1Ights of outputs 18 G Problems encountered and the1r solut10ns 26 H Plans for next year 27 I ProJect Performance Ind1cators 28 J Resource MobiiizatiOn 31 ProJect approved or on gomg 31 Proposal and concept notes 34 ~ 1 ENE 2005 1 ProJect SB-2 Conservatwn and Use of Tropical Genetlc Resources A PROJECT OVERVIEW ProJect Descnpt10n Objectlve To conserve the FAO Destgnated Collectwns and employ modem bwtechnology tools to tden!lfy and use gene!lc dtverstty for broademng the gene!lc base and mcreasmg the produc!lvity ofbean cassava nce and Brachwna Outputs 1 Improved charactenzatwn ofthe gene!lc dtverstty ofw1ld and cul!lvated spec1es and assoc1ated organ1sms 2 Genes and gene combmatwns used to broaden the gene!lc base 3 Increased efficiency ofNARS breedmg programs usmg bwtechnolog¡cal tools 4 Phaseolus Mamhot and forage spectes conserved mulllphed and d1stnbuted as per mtematwnal standards 5 Germplasm available restored and safely duphcated 6 Des1gnated Collectwns made soctally relevan! 7 NARS strengthened for conservatwn and use ofNeotropical plant genehc resources 8 Conservatwn of Des1gnated Collec!lons lmked With on farm conservatwn efforts and protected areas MIIestones 2005 Effic1ent transformation system developed for cassava Charactenzatwn of the bean core collectwns With 50 genom1cs and gene based microsatelhte markers Charactenza!lon of a bas1c collectwn of tepary bean germplasm w1th AFLP 1500 accessiOn from the cassava collec!lon genotypes With 36 SSRs markers Bean w1th h1gh 1ron and zmc tested and transferred to ClA T Afhca program for bwavaiiabihty testmg Survey of cassava germplasm for beta carotene SNP markers developed for bean and nce Targeted sequencmg of cassava geno me Molecular markers developed for dry matter content and res1stance to cassava green m1tes Isogemc ofQTL m nce developed and tested Gene expresswn stud1es for msect res1stance m Brachwna Differen!lally expressed genes for adaptatiOn of Brachzana to ac1d smls 1solated by microarray Bean cONA hbranes for drought generated Companson of gene flow m bean and nce under controlled and field conditiOns Technology for rap1d propagat1on system transferred to NARS 2 Testmg ofnce T DNA populatwns for gene tden!Ificatwn 2006 Scalmg up of marker asststed selec!Ion and gene!Ic transfonnatwn estabhshed for nce bean and cassava Marker asststed selected for mul!Iple tratts tmplemented m beans nce and cassava Target genes for drought tdentified and tested m beans Htgh tron and zmc bean !mes developed through markers asststed selectwn released for field testmg Beta carotene cassava tested m Colombta Braztl and selected countnes m Afnca Htgh protem cassava !mes developed and tested m Colombta and selected Afncan countnes Fteld testmg for transfonned cassava wtth Bt gene and transfonned nce wtth sheath bhght reststance Htgh through put propaga!Ion for selected troptcal frutts tm!Iated 2007 Allele mmmg of ex sztu/m sztu collectwns of wtld rela!tves of beans cassava for genes of economtc tmportance Gene flow studtes dtffused to NARS Candtdate genes for drought tolerance tdentified for bean and nce Gennplasm Upgradmg Plan completed Safety duphcates at CIMMYT and CIP for bean and cassava gennplasm Bwforttfied bean and cassava vanettes m field testmg Methods for raptd mul!tphca!ton of troptcal frutt gennplasm dtffused to NARS Fteld testmg for cassava transgemc !mes expressmg mductble flowenng genes for control of flowenng m cassava breedmg Users CIAT and NARS partners (pubhc and pnvate) mvolved m gennplasm conservatwn and crop genettc tmprovement and agrobwdtverstty conservatwn ARis from DCs and LDCs usmg CIAT technologtes B Prmc1pal Collaborators AfrzcaNARS DRC Mvuazt Research Center (INERA) Ghana Crop Research Instttute (CRI) Kumast Kenya Umverstty of Natrobt Malawi Clutedze Research Statwn Malawt N1gena Natwnal Root Crops Research Ins!ttute (NRCRI) Instttute for Agncultural Research and Tratmng (IAR&T) Ibadan Rwanda ISAR Tanzama Agncultural Research Instttute (ARI) Uganda Namunlonge Agncultural and Ammal Research Instttute Kampala Medtcal Bwtech Laboratones Kampala 3 Latm Amer1can NARS and Umversltles Bolivia CFP Centro Fitogenehco Pru.rurnam Brazii Embrapa Cenargen Embrapa CTA Embrapa CNPAF Embrapa CNPMF Umversity of Campmas Colombia Cemcana Cemcafe Umvers~dad Javenana CIB COLCIENCIAS Colombian Mmistry Agnculture and Rural Development Corp01ca Corporac10n Bwtech Colombian Nahonal Bwsafety Councii FEDARROZ ICA Instituto Humboldt UmAndes UmValle Umversidad Nacional at Palrmra and Bogota, Umversidad del Toiima Chile INIA REDBIO Costa Rica Umversity of Costa Rica Cuba INIVIT Domimcan Repubiic IDIAF National Bean Progrru.ns of the (INIAF) Ecuador INIAP Umversidad Catohca Honduras Zamorano Mexico Umversidad Autonoma de Mexico INIFAP Nicaragua Mimsteno de Agncultura Peru INIA Venezuela Centro Tecnologico Polar Simon Bohvar Umvers~ty Colomb1a NGOs CEGA FIDAR PBA REDBIO Colombia Latm Amenca Small Farmers from Pescador and Tierradentro Cauca Cauca farmers associatwn Parque del Software Cah Colomb1a pr1vate sector Com product Barranqmlla Agrobws Bogota LIMSYS Cah DATABIO Cah Syngenta, Cah As1aNARS Chma Academy of Agncultural Sciences (CAAS) SCIB India Central Tuber Crops Research Inshtute (CTCRI) Thiruvananthapurru.n Kerala Thadand Rayong Field Research Center Bwd1vers1ty Inst1tutes Colombia Instituto Humboldt Costa Rica Inb10 Mexico Conabw US Smithsoman Museum ofNatural History Advanced Research lnstltutes Australia Center for Apphed Molecular Bwlogy m Intematwnal Agnculture (CAMBIA) Europe Belgmm Umvers~ty of Ghent Denmark Umversity of Aarhus France CIRAD Genoplants IRD INRA Umvers1te de Perpignan Germany UmversJty of Fre1burg Umvers~ty of Hanover Umvers~ty of Hohenhe1m Federal Bwlog¡cal Research Centre for Agnculture and Forestry (BBA) Netherlands PRI Wagenmgen Sweden USLU Uppsala Switzerland Umvers1te de Geneve ETH UK Umverslty of Bath Japan JIRCA Tsukuba Umted S tates Clemson Umvers~ty Comell Umvers1ty Darlforth Center Kansas Sate Umvers~ty Lomsiana State Umverslty Michigan State Umverslty Natwnal Center for 4 Genome Research (NCGR) Ohw State UmversJty Penn State Umvers1ty Rutgers Umvers1ty Sm1thsoman Molecular Systematic Lab Umversity of Nebraska Umvers1ty of Puerto Rico Umvers¡ty of Ch1cago USDA Plant So!ls and Nutntwn Lab at Comell Umvers~ty USDA at Ch!ldren Hospital Baylor Umverslly USDA Soybean Genom1cs at Beltsv!lle Y ale Umvers1ty Regzonal networks ASARECA SACCAR AfNet ECABREN and SABRN (Afuca) SIGTT A (Central Amenca) REDBIO (Latm Amenca) CATIE and EAP Zamorano (Central Amenca) Cassava Bwtechnology Network (CBN LAC) FLAR CLAYUCA CGIAR and lnternatzonal orgamzatzons CIP CIMMYT FAO IAEA ICARDA ICRISAT IFPRI UTA IPGRI IRRI TSBF WARDA CGIAR Challenge Programs HarvestPius Generation CGIAR system hnkages Savmg Bwdiversity (40%) Enhancement & Breedmg (55%) Trammg (4%) Informatwn (!%) CIA T proJect hnkages lnputs to SB 2 Germplasm accesswns from the gene bank proJect Segregatmg populatwns from crop product!Vlty proJects Charactenzed msect and pathogen strams and populatwns from crop protectwn proJects GIS serv1ces from the Land Use ProJect Outputsfrom SB 2 Management ofDesignated Collectwns (gene banks) genetic and molecular techn1ques for the gene bank crop productivity and s01ls (m¡crobial) proJects Identified genes and gene combmatwns for crop product!Vlty and protecl!on proJects Propaga!Ion and conservatwn methods and techn¡ques for gene banks and crop product!Vlty proJects Interspecific hybnds and transgemc stocks for crop producl!vity and IPM proJects 5 C Budget BUDGET 2004 PROJECT SB2 Conservabon and Use of Trop1cal Genet1c Resources SO URCE AMOUNTUS$ PROPORTION (!) Unrestncted Core 599 902 14/ Restncted Core a 01 Carry over from 2003 493 68f 121 Sub total 1 093,591 26/ Spec1al Pro]ects 3 061 200 721 Generat1on Challenoe Prooram 125 000 3/o [rotal Project 4 279 791 100/ *The largest part of the carry o ver funds ts related to USAID ProJects wtth CIA T partners m process of defimtton HARVESTPLUSBUDGET BUDGET 2004 SOURCE AMOUNTUS$ PROPORTION (1) Unrestncted Core o 0/ Restncted Core o 01 Carrv over from 2003 o 01 Sub total o 0/ Restncted Pro]ects 1 1 550 000 100/ h"otal ProJect 1 550 000 100/ 1 lncludes only funds 1mplemented by CIAT 6 D CIAT SB-2 ProJect Log Frame (2005 2007) PROJECT CONSERVA TJON ANO USE OF TROPICAL GENETIC RESOURCES PROJECT MANAGER JOE TOHME NarratJve Summary Measurable Indtcators Means of Venficat10u Importan! Assumptions Goal To contnbute to the # of CIA T sctentlsts and CIAT and NARS pubhcatwns sustamable mcrease of partners usmg b10technology Statlstlcs on germplasm productlVlty and quahty of mformatlon and tools m crop exchange mandated and other pnonty research crops and to the conservat10n Germplasm and Genetlc stocks of agrobtodtversily m troptcal available to key CIA T partners countnes Purpose To conserve the genetlc A database on dtverstty ofwlld Pubhcatlons reports and Pro active parttctpatiOn of dtvemty and ensure that and cultivated spectes proJect proposals CIA T and NARS charactenzed Mapped economtc genes and agncultural sctentlsts and agrobtodtverstty tmproved gene complexes biOlogtsts crop genettc stocks and Improved genetlc stocks !mes modern molecular and and popula!Ions cellular methods and tools are used by CIAT and NARS sctenttsts for tmprovmg usmg and conservmg crop genetlc resources Output 1 Genomes charactenzed of Molecular Genetzc Technzques wtld and culttvated spectes of and molecular znformatwn on bean cassava nce and dzverszty Brachtana and of assoctated organtsms 2005 Method avatlable Pubhcatton Avatlabthty of up to date SNP markers for bean and nce of SNP pnmers and protocols genomtcs eqmpment Development of genome developed Reports on marker anal vsts and Collaborat10n wtth NARS 7 Narrative Summary Measurable Ind1cators Means of V enficat10n lmportant Assumptlons w1de anchored PCR based artlcles descnbmg the genetlc mamtamed and expanded markers for marker ass1sted Charactenzat10n of core structure of the bean and selectlon gennplasm collect10n and natlonal cassava core collect10n of the charactenzatlon and fine collect10ns ofbean w1th fifty world gennplasm collect10n mappmg and gene flow genom1c and gene based VIS a VIS other acceSSIOnS ofthe analySIS m!Crosatelhte markers collectlon Identlficat10n and mappmg Charactenzat10n of a bas1c of use fu! genes and gene collect10n of tepary bean combmat10ns for gennplasm w1th AFLP and agronom1cal and nutntlonal m1crosatelhte markers Ava1lab1hty of a laboratory tra1ts mfonnat10n management 1500 cassava access10ns system (LIMS) Markers ass1sted select1on genotyped w1th 36 SSR Reports and pnmers made for mulhple tra1ts for bean markers ava¡lable m databases cassava and nce Genes smtable for the B10mfonnatlcs tools for data Cassava core collectJOn development of COS markers mmmg screened for carotenes and true m cassava databases retentlon of carotenes after processmg detennmed Art1cles descnbmg molecular LIMS developed and markers assoc1ated With Implemented agronomJcal tra1ts 2006 07 Populahons markers and BAC Cross legume and smgle hbrary ava1lable for dJstnbutJOn nucleot1de polymorphism and further analys1s by partners markers m CIA T mappmg populat10n mtegratlon of legumes cross collect10n dJversJty data ------~ -------- 8 Narrattve Summary Measurable Ind1cators Means of Venficatwn Important Assumptlons Selecl!on of a set of genes for the development of cassava COS markers Pubhcatwns reports and data Jdent¡ficatwn and mappmg on populatwn posted on web useful genes and QTLs for Databases shared agronomzcaltralls m bean cassava rzce and Brachzarza Map posJI!on softarget genes md1cated 2005 QTL analys1s completed m two bean populatwns for nutntwnal tra1ts mcludmg 1ron zmc and tarmm content Molecular markers tlghtly hnked to CMD res1stance 1denhfied and BAC hbrary of TME3 constructed Two advanced backcross w1th w1ld AA Oryza genomes genotyped and QTLs for y¡eld componen! m nce 1denhfied Generatwn ofnce mappmg populahons for nutnhonal tra1ts mcludmg 1ron and zmc 2006 07 QTL analys1s completed on - 9 NarratJve Summary Measurable lnd1cators Means of VenficatJon lmportant Assumpt10ns two populatwns of common bean for drought tra1ts and nutnhonal tralts One advanced backcross populatwns (BC2F5) from wlld beans 1s genotyped to determme 1f nutnhonally supenor genotypes can be obtamed lntegratwn of drought and nutntlon QTLs across mulhple populatwns Populatwns segregatmg for dry matter cyanogemc glucos1des content leaf retentwn res1stance to homworm developed charactenzed and evaluated w1th molecular markers QTL analys1s completed on one populat10n for Al tolerance m Brachwna Output 2 Genomes modlfied genes Candzdate genes zdentzfied for and gene combmatwns used agronomzca/ trazts to broaden the genehc base of Phenotyp1c bwchem1cal crops (bean nce and 2005 analys1s conducted pnor to cassava) and forage spec1es Clomng of cand1date genes Pubhcatwns reports and molecular analys1s 10 Narrat1ve Summary Measurable Ind1cators Means of Venficahon Important Assumpt10ns . (Brachwna) mvolved m tolerance lo ac1d proJect proposals 1 smls fulllength cDNA IdenllficatiOn of pomts of hbranes developed Germplasm L1branes and genellc mtervent10n and cand1date genes avaiiable . mechanism of plan! stress Differenllally expressed genes mteract10n Isolated by microarray Libran es and candidate genes avmlable Improved methods for Molecular charactenzatiOn of genellc transformallon for spittlebug msect resistance m Libranes and candidate genes bean nce and cassava Brachiana usmg cDNA ava!lable subtrac!Ive differenllal Develop and obtam gene expressed hbranes constructs for traiis related to plan! disease msect Comparallve genomics and Microarray chips avmlable for resislance plan! stress and gene discovery drought m bean distnbutiOn Contmued access lo nutnllonal traits by gene express10n profihng b10safety field testmg and cDNA hbranes produced under collaboratiOn WIIh CIRAD Acquisiilon ofnce T DNA drought conditiOns IRD and genoplantes and Ac/Ds mutants Charactenzat10n data made populallons for testmg and Cassava bactenal bhght avmlable on web Databases gene discovery mteract10n charactenzed usmg Phenolypic biochemical cassava cDNA plan! defense anal ysis conducted pnor lo Irnplement b10safety microarray chip of 6000 molecular analysis 1 regulallon for greenhouse cassava umgene sets Libran es and sequence made and field cond!IIOn avaiiable Databases T DNA nce 10000 mutants collectiOns charactenzed under Fundmg from Rockefeller field condiilon Techmcal reports on si les and Foundat10n access to distnbutiOn ofwiidlweedy/ genes IPR management to 2006 07 landraces and cases of gene access genes and gene Consohdated genes sequence flow promoters B10safety data for drought and acid smls regulat10ns m place 1 stress response pathways gene -- - - --- 11 Narrat1ve Summary Measurable lndtcators Means of Venficat10n Important AssumptJons chtps for candtdate genes developed over 5000 genes Number of advanced !mes and arrayed and 25 candtdate genes crosses w1th RHBV res1stant putatlvely related to drought transgemc source tolerance Jdentlfied IPR management to access Modtficatlon of flowenng to genes and gene promoters mcrease the effic1ency of Btosafety regulatwns m breedmg m cassava by usmg place constructs for LFY AP 1 and Request extensmn of field FLC flowenng genes under the permtt to other field control of an ethanol mducJble locatlons m Colombia promoter approved Clomng of the cassava mosa1c IPR management to access dtsease (CMD) reststance gene genes and gene promoters CMD2 by posJtlonal clomng Bmsafety regulatwns m Pubhcatlons reports and place protocols made avatlable and Transformatzon pubhshed 2005 Airead y recetved approval Degree of gene flow from nce for field testmg F1eld and bean mto wild/weedy Bwsafety mspected and relatlves measured m trop1cal approved centers of diversJty Identtficatmn evaluat10n and d¡ffus¡on of molecular markers for assessmg rate and dtrectlon ofgene flow 12 1 Narratlve Summary Measurable lnd1cators Means of Venficahon Important Assumpt10ns F1eld performance bwsafety evaluatwn for agronom1c tra1ts of advanced generatwns of crosses between RHBV N res1stant transgemc nce and selected nce commercml vanetles Transgemc Agrobacterzum strams generated w1th constructs from JIRCAS contammg d1fferent verswns of DREB gene encodmg for drought tolerance Transgemc !mes of cassava w1th Bt constructs and or nce w1th res1stance to sheath bhght tested under bwsafety field cond1t1ons and evaluated for agronom•cal tralts 2006 07 Protocol developed for generatmg transgemc plants based on mannose selectwn system m place for nce and cassava Scalmg up ofnce and cassava transformatwn effic1enc1es mcorporatmg new genes cultlvars and regeneratlon - 13 Narrative Summary Measurable lndJcators Means of VenficatJon Importan! Assumptwns methods and testmg plants m the field F1eld testmg for nce and cassava transfonned genotypes Ophmizahon of bean transfonnat10n protocols Ophmizahon of low cost altemahve system usmg temporary ImmersJOn system pnnc1ple m place for nce anther culture callos mductJon and plant regenerat10n for a scalmg up system Adaptahon and ophmizahon of protocols for nce 1solated m1crospore culture as an altemahve for h1gh effic1ency generahon of doubled hapl01ds Output 3 Increased effic1ency ofNARS 2005 breedmg programs usmg Marker ass1sted select10n for PubhcatJOns Govemment and mdustry bJOtechnological tools Cassava Mosmc D1sease Trammg courses and support nat10nal b10tech (CMD) transferred to NARS m workshops ImhatJves Tanzan1a Pro J ect proposals CIA T partners m LDCs usmg mfonnat10n and genehc stocks LAC NARS mvolved m RegiOnal workshop 14 NarratiVe Summary Measurable Ind•cators Means of Venficat10n Important Assumpt10ns b10fortlficallon effort for 1ron zmc and beta carotene Improved capac1ty of MT A estabhshed and JOmt Freedom to operate Colomb1an NARS to deal w1th pubhcat10n obtamed b10safety 2006 07 New partnersh1ps w1th pnvate sector Agreement on technology and gene constructs access 15 NarratJve Summary Measurable ludicators Meaus ofVenficatiOn Important Assumpt10ns Output 4 Bean Cassava and forage 2005 spec1es conserved and GermmatJon rates for long V1s1ts to GRU substatJons and Absence of uncontrolled multJphed as per mtematlonal stored matenals conservatlon facJhtles d1seases standards Cost per accessiOnlyear Quarantme greenhouse compared w1th other gene space ava1lable at d1fferent banks altitudes 2006 07 Safety duphcates at CIMMYT and CIP for bean and cassava ' germplasm Narratlve Summary Measurable lndicators Means ofVenficatiOn Important Assumptlons OutputS 2005 Germplasm available Number of germplasm requests VJsJts to multJphcatJOn plots Agreement w1th CIA T restored and safely rece1ved and fulfilled annually Reports on requests and holds duphcated dehvery CIA T becomes partner to Low cost rap1d propagat10n Number of core collectJOns the Treaty system for cassava multJphed and sh1pped 1mplemented w1th farmer assocJatJon Users rece1ved germplasm and data 2006 07 Cryo conservatlon technology developed tested and 1mplemented for cassava Users asked for novel 16 Narratlve Summary Measurable lndicators Means of Venficatwn Important Assumptwns ! germplasm and data Output6 2006 07 Des1gnated Collectlons made Landrace dJversity restored to Germplasm catalogs IntematiOnal collectmg socially relevan! farmers Plan! vanety registratiOn logs poss1ble Farmers use new van elles Nat10nal catalogs Quarantme matters cleared Breeders use novel genes Output 7 Strengthen NARS for 2005 conservaban and use of NARS germplasm collect10ns Country questwnnmres NARS and networks Neotrop1cal plan! genetlc conserved Courses reg1stered wiihng to cooperate resources Number oftramees tramed at DistnbutiOn and sales of CIAT trammg matenals 2006 07 Methods for rap1d Protocols pubhshed multlphcatiOn of selected tropical frmt germplasm d1ffused to NARS Number ofumvers1t1es and NARS usmg trammg matenals Output 8 Conservatlon ofDes¡gnated Number of case stud1es and ProJect documentatlon NARS mterested m Collecbons hnked With on p1lot m sllu conservatlon Databases conservaban efforts farm conservat10n efforts and proJects Farmers mterested m , protected are as conservat10n efforts 17 E Current SB-2 InvestJgators Dtsctphne, posttton and nme frachon T1rne N ame DISCipline ded1cahon % Al ves Alfredo CBN RegiOnal Coordmator 100 Beebe Steve Bean Breedmg 30 BellottJ Anthonv Cassava Entomology 20 Bla1r Mathew Bean Genetics and breedmg 70 Ceballos Heman Cassava Breedmg 40 Chavamaga, Paul Transgenesis Cassava 100 Debouck Daniel Botan y 20 Fregene Martm Cassava Genetics and 60 breedmg Ishitan¡ Manabu Molecular B10log¡st 100 Lentmt Zatda BIOlogy/Genettcs 80 Loneux Mathtas Rice Genehcs 50 Martmez Cesar Breedmg 49 MeJta Alvaro Cell B10logy 100 Sperlmg Lomse Seed Systems 20 Tohme Joe Genomtcs Pro)ect Manager 100 F Htghhghts of outputs Staff changes Dr Valene Verdter IRD wtll be spendmg 3 months at CIAT to work onJomt proJects related to nce -pathogen mteractlon Dr Verdter s stay wtth the proJect to remforce the collaborat10n between CIAT and IRD and wtll allow the team to prepare JOmt proJects for the Generatlon Challenge Program Dr Verdier w!ll work closely wtth Fernando Correa Mathias Loneux Cesar Martmez and Joe Tohme Team member rece1ved the followmg awards • Gallego Gerardo PhD 2004 EstudiOs prehmmares para el atslarruento y clonac10n de genes de reststencta a pyrzculana grzsea en arroz usando la estrategia de clonac10n posiciOnal Uruverstdad NaciOnal de Colombia Sede Palmtra. Mentona • Gattan - Sohs Gmtan PhD 2004 Obtenc10n y uso de secuencias rrucrosatehtales GAICA en estudiOs de diversidad genettca en las espectes de palmas colombtanas Atta/ea amygdalma Ceroxylon alpmum y Ceroxylon sasazmae realizada en la Uruverstdad NaciOnal de Colombia, Palmtra Mentona 18 • Arango Adnana MSc 2004 Identlficatlon of cand1date genes for alummmm res1stance m Brachwna Umvers1dad Nacwnal Bogota Mentona • Gonzalez Torres R I MSc 2004 St1mat10n of gene flow on Phaseolus vulgans usmg molecular markers m1crosatelhtes and polymorph1ms of chloroplast Thes1s awarded w¡th Ment recogn¡tJon by UDJversJdad Nac10nal de Colombia • Beltran Jesus Prem1o NaciOnal Hemando Patmo AsocJacJOn Colomb~ana de Ctenctas Bwlog1cas MeJor TrabaJO de Pre grado Tnges1mo cuarto Congreso Nacwnal de la Asoctacwn de Ctenctas Bwlog¡cas !bague Tohma Oct 12 15 04 • Soto Suarez Maunc10 Lopez Camilo Restrepo Stlvta P1egu Benolt Cooke Richard Delseny Mtchel Tohrne Joe and Verdter Valene Anahsts de expreswn global durante la mteracc10n yuca Xanthomonas axonopodzs pv mamhotzs usando mtcroarreglos de ADN Best work selected dunng the 11 Congreso NaciOnal de BJotecnolog•a Bogota • Gonzalez Torres R I E Galtan M C Duque O Toro J Tohrne & D G Debouck 2004 Measure of gene flow on Common bean usmg molecular markers F1rst Award for best poster Memonas 11 Congreso Colombtano de 8JOtecnologJa Bogota Colomb~a 210 211 • Gonzalez Ehana Fory LUJsa Fernanda Pmeda Rosana Rutz Paola Vasquez Juan Jose Corredor Edgar Duque Mynam Cnstma Stlva James y Lentlm Zatda Caractenzacton de la Dtvers1dad Genetlca Morfolog1ca y Fenologtca de Cuatro Poblacwnes de Arroz Maleza (Arroz RoJo) Colectadas en las Regwnes de HUlla y Tohma Second best poster at 11 Congreso NaciOnal de 810tecnolog•a- Bogotá Overall vtew of the SB2 project The team members lead successfully the use and mtegratlon w1thm CIA T of bwtechnology w1th breedmg actlvttles and the conservatlon charactenzatwn of gen elle dJVerstty to 1) Irnprove the nutntwnal quahty of crops aclueve and sustam a contmuous y¡eld mcrease to meet the food needs of a rap1dly growmg populat10n 2) Conserve the natural resources base needed for future development 3) Irnprove the hvehhood of the rural poor who have not benefited so far from the technologtcal advances To achteve tts goals the team members have mteracted w1th researchers from Advanced Research Instttute Natwnal Programs NGO and the pnvate sector They also pursued successfully al! three outputs related to Output 1 Output 2 Output 3 Irnproved charactenzatwn of the genetlc dtverstty of wild and culttvated spectes and assoctated organ1sms Genes and gene combmatlons used to broaden the genetlc base Increased effictency ofNARS breedmg programs usmg bwtechnolog1cal too1s 19 Only the followmg aclnevements are summanzed Measunng Genetlc D1vers1ty ID Common Bean The m1crosatelhte markers we have developed at CIA T have g¡ven us the opporturuty to d1ssect genehc d!vers1ty m common bean on a larger scale and to a finer degree than has ever been poss1ble before D1vers1ty was analyzed m a two collectwn of common bean accesswns from Colombia and the Canbbean as well as m tepary bean accesswns Both Colombia and the Canbbean s1t at cross roads of gene flow between the centers of ong¡n for common bean Therefore many Andean genotypes from these reg¡ons show poss1ble mtrogresswn of Mesoamencan alleles We wanted to document th1s phenomenon as a bas1s for further genotypmg of the Andean and Mesoamencan core collechons as well as natwnal collechons from Bolivia Braz¡J Chma Colombia and Mex1co wh1ch we are domg as part of the Generatwn Challenge Program Tlns has mvolved the development of new tools for evaluatmg genotyp1c d!vers1ty G1ven that m¡crosatelhtes are the most rehable polymorph!c marker system we are concentratmg on developmg panels of fluorescent common bean m1crosatelhtes useful for genotypmg We ha ve now determmed the d1scnmmahon power and allehc d¡vers1ty values for all m1crosatelhte markers developed at CIA T and plan to use th1s mformatwn m further genotypmg and assoc1ahon mappmg stud1es Study of bean gene flow Phase 1 of the Gene Flow ProJect supported by BMZ of Germany has shown that m the bean model m Costa Rica gene flow between the crop and 1ts wild relah ve occurs though preferent~ally from the wild form mto the culhvated Intermed1ate forms hypotes1zed to result from gene flow events were proven to be so Flow from the culhvated mto the wild !S low but s1gmficant and occurs repeatedly m space and time The results were obtamed through molecular markers based on both nuclear and cpDNA genes Gene flow on statwn m Costa Rica usmg commerc¡aJ !mes was demonstrated to be low usually below 1% Ways of computmg the data seem s1gmficant however w1th lngher percentages when one computes the number ofhybnd plants mstead ofhybnd seeds Th1s could explam many d1screpanc¡es m outcrossmg rates found m the hterature In farmers plots m Costa Rica a lngher outcrossmg of 6 8% was found when usmg landraces as compared to commerc!al !mes An 1mage analys1s of shgma areas between w¡Jd forms tradltwnallandraces and modero culhvars has shown a larger termmal area m the Wild wh1Ie culhvated forrns ha ve a b1gger mternal area Th1s could explam many figures m populatwn genehcs ofthe crop model ExpandiDg the Scope of Marker Ass1sted Selectlon ID Common Bean at CIAT Marker ass1sted selectwn 1S a pnonty m the bean breedmg program because of the large number of segregants screened and the d1verse array of b1ohc and ab1ohc hm1tahons bemg tackled Unhl recently all marker ass1sted select10n m common beans was done w1th SCAR marker In the past year we have begun to 1mplement severa! gene spec1fic m1crosatelhte markers for the selectwn of hnked trruts We have tested m1crosatelhtes for selechon of a rutrogen fixahon QTL m populatwns denved from BAT477 (also and 1mportant drought tolerance so urce) and for selectwn of bruclnd res1stance based on the Arcehn res1stance gene 20 In addJtlon we contmue to vahdate additiOnal SCAR markers m practica! real hfe plan! breedmg Situatlons We ha ve now made over 4000 plan! selectwns for the SCAR markers for the be 3 and dommant J BCMV res1stance genes and for the Co 42 and Co 5 anthracnose res1stance genes The be 3 marker was found to work well m Andean breedmg !mes especmlly for screenmg of chmbmg beans where 1! resulted m substantlal savmgs m plantmg area and agronom1c effort Meanwh!le the anthracnose markers were Improved for screemng w¡th alkahne lys1s extracted DNA All of th1s work bmlds on las! year s successes m usmg SCAR markers for bgm 1 and for QTLs for resistance to bean golden yellow mosa1c virus (BGYMV) and common bactenal bhght (CBB) Next year we plan to Implement additwnal microsatelhte and SCAR markers for addJtwnal res1stance genes and abwtlc stress QTLs QTL mappmg of nutntJOnal quahty m common bean QTL mappmg ofnutntwnal tra1ts has become an Importan! way to mcrease our understandmg of how to breed common bean for better mmeral content as part of the Harvest Plus Challenge Prograrn We are usmg a m1x of traditwnal bwchemistry genetlcs and genom1cs to d1ssect nutntwnal quahty traJts Collaboratwns wJth USDA Houston IS centenng on a bas1c mechamsm for mmeral uptake m legumes usmg common bean as a model for the tropical legumes As part of the overall genom1cs approach mformatwn from other well stud1ed spec1es such Medzeago truneatula peas and soybeans as well as other model spec1es such as Arabzdopszs thalzana wh1ch have extens1ve genetic and molecular resources are bemg used for gene d1scovery and func!Ional analys¡s A collaboratwn with the Grusak lab at USDA Houston 1s producmg very mterestmg results on the analysis of 1ron reductase as a mechamsm for enhanced 1ron uptake m common beans So far there IS ev1dence that there are d1fferences between parents of severa! mappmg populations for the1r ab1hty to reduce 1ron and that these d1fferences are ev1dent more at low Fe concentratwn !han at h1gh Iron concentratwn QTLs for 1ron reductase actlvity are located m the sarne genom1c regwns as sorne for uon accumulatwn More mformatwn on th1s trmt will be reported next year when severa! sets of recombmant mbred !mes have been fully tested and the QTLs for th1s trmt defim!Ively locahzed In addJtlon to mcreasmg 1ron content per se we are studymg the 1nhentance of soluble and msoluble tarmms wh1ch are Irnphcated as anti nutnents m mmeral absorpt10n through a QTL analys¡s of parents and segregatmg populatwns of common bean We also refined the gene!Jc map for the mmeral accumula!Jon mappmg populatwns and added phenotyp1c data showmg pattems of genotype x environment mteractwn This bmlds on our research reported last year on the QTL analys1s of mm eral content ( espec1ally of 1ron and zmc accumulatwn) All of th1s research forms part of the Harvest Plus challenge prograrn Molecular marker ass1sted farmer partlpatory breedmg proJect to 1mprove local cassava vanet1es m Tanzama with resistan ce to pest and d1seases The Tanzan1an MAS proJect funded by the Rockefeller founda!Ion seeks to transfer useful vanabihty from the crop s center of diversily of cassava to Afnca This year a total of 335 BC2 progerues (AR hnes) that combme res1stance to CMD and to the cassava green mites (CGM) denved from a w!ld rela!Ive and 207 genotypes (CR !mes) obtmned from crossmg CIAT ehte parents and CMD resistan! !mes were mtroduced from Colombia to Tanzama m three sh1pments The plants were hardened m the screen house evaluated for frog skm disease (FSD) and then transferred to the field They w!ll be evaluated later tlus season and 21 no less than 60 genotypes selected based on evaluat10n of h1ghly hentable agronom1c traJts for crosses to 90 local vanetJes selected from all over the country Molecular markers assoc1ated w1th CMD and CGM w!ll be used to d1scard much of the resultmg segregatmg populat10ns so that the breeder and farmers can concentrate on a small number of progeny havmg res1stance to the pnnc1pal pest and d1sease and farmer/end user preferred tra1ts The concept of the Tanzaruan MAS proJect JS already bemg extended to addJt!Onal NARs m Afnca, the AR and CR !mes have been sh!pped to Uganda and N1gena already m preparat10n for crosses to local vanet1es ldentificatwn of sources delayed post harvest physwlogJcal detenoratlon (PPD), res1stance to whJteflJes and hornworms m wdd Ma01hot spec1es and development of populatlons to study genet1c mhentance and for mtrogresswn of the tra1ts mto cassava Post harvest phystolog¡cal detenorat10n (PPD) and anthropod pests are severe marketmg and productlon constramts respectlvely m cassava lt has been esttmated that cassava farmers typtcally resource poor farmers lose 48 mtlhon tons of fresh root valued at US$1 4btlhon every year to pests dtseases and PPD sorne 30% of total world product10n Dramatlcally delayed PPD has been tdentJfied m mter spec1fic hybnds from Manzhot walkerae The delayed PPD tratt ong¡nally from the wtld Manzhot paren! was successfully transferred to an F1 mter spec1fic hybnd suggestmg a dommant or addttlve gene actJon of gene(s) mvolved The only source of reststance to the cassava homworm was also 1dentJfied m 4th backcross denvatJves of M glazwv¡¡ Moderate to h!gh levels of reststance to whtte fltes have also been found m mter spectfic hybnds of M esculenta sub spp jlabellifolza Agam reststance was recovered eastly m F 1 mter spectfic hybnds suggestmg a stmple mhentance of the tratt Etght mappmg populatJons have been developed for marker asststed study of the mhentance ofthese tratts Functwnal Genom1c Tools to Study Starch Content and Cassava Bactenal Bhght Res1stance Two economtcally tmportant characters starch content and bactenal bhght reststance were targeted to generate a large collect10n of ESTs and Mtcroarray analysts For ESTs collectJon two hbranes were constructed from cassava root tlssues of vanetJes wtth htgh and low starch contents Other hbranes were constructed from plant tlssues chaJienged by the pathogen Xanthomonas axonopod1s pv mamhot1s (Xam) We obtamed 11 954 cONA sequences from the 5 ends mcludmg 111 from the 3 ends Cluster analysts permttted the tdentJficatJon of a umgene set of 5 700 sequences Sequence analyses perm1tted the ass1gnment of a putatJve funct10nal category for 37% of sequences whereas -16% sequences d1d not show any s1gmficant Slm!lanty w1th other protems present m the database and therefore can be cons1dered as cassava spectfic genes A cassava cONA m1croarray contammg the un1gene set was constructed and used to study the mcompatJble mteractlon between cassava and Xam A total of 199 genes were found as d¡fferentJally expressed (126 up regulated and 73 down regulated) A greater proport1on of genes d1fferentJally expressed was observed at 7 days after moculat10n Express10n profihng and cluster analyses md1cate that m response to moculatJon W1th Xam cassava mduces severaJ genes mcludmg pnnc1pally those mvolved m ox1datJve burst protem degradatlon and 22 pathogenests related (PR) genes In contras! genes encodmg protems that are mvolved m photosynthests and metabohsm were down regulated In addttlon severa! other genes encodmg protems wtth unknown funct10n or showmg no stmtlanty to other protems were also mduced The QRT PCR expenments allowed to confirm the rehabthty of our mtcroarray data In addthon we showed that sorne genes are mduced more raptdly m the reststant than m the susceptible cultivar Thts ts the first large cassava EST resource and umgene mtcroarray developed today and pubhcly avatlable thus makmg a stgmficant contnbut10n to genomtc knowledge of cassava Cassava genettc modtficatwn - Bwsafety F1eld testmg The first transgemc cassava plants produced m a CG center have been planted m an open field of CIA T The government of Colombta gave permtsswn on August 2004 to plant about 100 p1ants of three clones that carry a gene for msect reSIStan ce These plants wtlltmtlally be grown to multlply the seed (cuttmgs) to perform tnals for agronomtc evaluatlon As a precautlon to av01d gene flow wtthm CIA T campus plants wtll not be allowed to flower Bestdes the cassava germplasm collectlon at ClAT ts not planted m the field m 2004 2005 so there are very httle chances of transgenes movmg to the cassava collect10n m the field The transformatlon effictency of cassava can double by modtfymg the temperature at whtch Agrobactenum msert thetr genes mto plant cells A proJect financed by the USAID whtch ts part of collaborat10n wtth Ohto State Umverstty atmed at mcreasmg the transformat10n effictency of cassava The results obtamed so far mdtcate that wtth lower co culture temperatures twtce as many plants can be produced m a smgle transformat10n expenment Molecular phenotypmg for crop tmprovement under abwlic stresses The strategy we have been takmg to tmprove crop performance under abtotlc stress condttlons ts to 1) tntegrate phys10logtcal aspects on tratts of mterest usmg dtverse genetlc matenals that come out from breedmg actlvtty pnor to apphcat10n of molecular tools 2) lmk obtamed molecular components to breedmg (e g marker development) 3) expand genomtc tools such as full length cDNA clones to facthtate molecular too! development for breedmg We focused on tdentlfymg key molecular components m cornmon bean to mcrease plan! performance under water hmtted condtt10ns Th1s acttvtty was m part supported by Generatlon Challenge Program Thts mvolves 1) tsolat10n of the genes mvolved m response to water stress 2) evaluatton of gene expresston m relatton to phenotyptc tratts (e g deeper root) and 3) development of useful molecular markers to select genotypes wtth better performance under water stress condthons We selected 100 to 200 genes from spectes such as Arabtdopsts representmg twelve funct10nal categones based on pnor evtdence suggestmg these genes are mvolved m stress response or metabohc pathways In collaborat10n wtth JIRCAS Japan Drought Responstve Element Bmdmg (DREB) gene was cloned from the bean and the gene expresswn analysts ts under way usmg breedmg matenals (e g BAT477 and G21212) that was shown dtstmct phenotypes (deeper rootmg and htgh mobthzatwn of photosynthates) under drought stress 23 Gene Flow The first phase of a 4 year BMZ funded proJect on the assessment of gene flow from crop to w¡ld/weedy relatlves m trop1cal Amenca usmg bean and nce as models was accomphshed The goal of tlus proJect !S to generate basehne genetlc mformat10n for the development of gmdehnes on the safe mtroduct10n and use of novel agnculture trruts (b10technology den ved or not natlve from the place of mtroduct10n) while reducmg potentlal env1ronmental 1mpact on natlve b!Od!vers¡ty m the Neotrop1cs The model crops selected are 1mportant for food secunty throughout Latm Amenca Afnca and As1a bemg bas1c for the rural and urban poor people Results showed (deta1led mformat10n are found m SB2 Reports from 2001 to 2004) that on both crop models (beans and nce) gene flow occurred 1) between the crop and 1ts w¡ld/weedy relatlves u) the d1rect10n from the crop to the Wild/weedy relatlve cannot be 1gnored (from O 003% toO 06%) and m) mtrogress10n because of gene flow 1s repet1t1ve m space and lime (on the bean model m nce repeated mtrogress10n over lime needs to be evaluated) The sconng of phenotyp1c tra1ts alone (1 e herb!C!de res1stance anthocyanm color m flower/stem/leaves) to assess gene flow overestlmates the level of hybnd¡zat!On rate Because of these potentlal errors methodolog¡es were optlm1zed to use molecular markers to detect rate and d1rect10n of gene flow m bulk DNA allowmg the analys1s of large field samples These tools and methodolog1es generated w¡Il g1ve a better understandmg of the gene flow/mtrogress10n dynam1cs m crop/w¡ldfweedy complexes and potentlal 1mpact on b10d1vers¡ty The potentlal 1mpact of gene flow on the genetlc structure ofthe rec1p1ent populat10n stlll needs to be assessed To that end we are plannmg m future stud1es to use genes that confer a pos¡tlve advantage agamst select10n also allowed estabhshmg a methodology to detect cases of mtrogress10n at small scale m crop/weedy/w¡ld contact zones and to quantlfy 1t w1th the a1d of molecular markers B1osafety Capac¡ty Buildmg The heavy mvolvement of umvers1ty departments and b10safety authontles !S key to the mstltut10nal strengthemng and the development of sc1ence based pohc1es on b10safety m developmg countnes Towards that end CIAT 1s playmg an active role on env1ronmental b10safety research conducted m the Neotrop1cs and on capaclty bmldmg As part of a 4 year proJect financed by BMZ (Germany) CIAT conducted a workshop on gene flow analys1s and 1ts 1mphcat10ns bemg for the conservat10n of landraces m sztu or the mtroduct10n and management of modero !IDproved cultlvars A total of 41 partiC!pants of Umvers1t1es and pubhc Inst!tut10ns respons1ble for 1mplementmg b10safety regulat10ns from Mesoamenca Central Amenca, Andean reg¡on and Braz¡J attende The part¡c¡pants mcluded the areas w1th h1ghest biOd!Versity of th1s reg¡on and w1th d1fferent leve! of development and expert1se on bwsafety Because of CIA T mvolvement on environmental bwsafety research 1t was mvlted to g¡ve support toa GEF World Bank proJect approved for Colomb¡a to 1mplement the Cartagena Protocol and as part of tlus Imlitallve CIAT was requested to offer a senes of courses to bmld techmcal capac1ty for the analys1s of GMOs and of potenllal envlronmentaJ¡mpact The first B10safety Workshop for the Colomb1an Inter Ins!itullonal Group respons1ble for Implementmg the Cartagena Protocol was offered on October 4 6 2004 and mcluded a total of 31 partiC!pants from the Mm1senes of Agnculture Health Env1ronment Cornmerce Fore1gn AffalfS Inv1ma ICA Inslltute Von Humboldt CVC Nallonal Department ofPlannmg Colomb1an Natwnal Department ofSc1ence Agnculture and Animal Natwnal Techn1cal Bwsafety Cornmlttees 24 CBN s ActiVIties for 2004 The Cassava Bwtechnology Network for Latm Amenca and the Canbbean (CBN LAC) 1s a network of cassava researchers and end users rummg to enable small scale cassava farmers processors and consumers to benefit from advances m cassava bwtechnology enhancmg the val u e of cassava for food secunty and econom1c development m the poorest rural areas of the LAC In th1s fouth year (2004) of operat10n the CBN LAC devoted to the followmg mam acl!villes a) Implementmg momtonng and gUidance of 11 proJects (pilot s1tes and small grants programs) m Colombia Braz!l Cuba and Ecuador b) Scholarsh1ps for postgraduate stud1es m biOdiversity under the Gmes Mera Memonal Fellowsh1p Fund e) Reahzal!on of the S1xth lntemal!onal CBN meetmg (CBN VI) The mam ach1evements of the CBN s proJects were 1) The establishment of propagal!on systems at farm leve! for both low cost m vztro m•cropropagatwn and rap1d propagal!on systems wh1ch have allowed farmers to clean local and 1mproved vanel!es of d1seases and pests ensunng the1r food secunty and preventmg the loss of diver51ty and 2) bUI!dmg capac1ty for natwnal cassava R&D programs sl!mulatmg collaboral!ve research proJects between less developed countnes and advanced laboratones through CBN small grants program The CBN VI held at CIAT from 8 to 14 March 2004 hosted 148 participants from 28 countnes from all contments The theme of the meetmg was Addzng val u e to a small farmer crop to d1scuss how biOtechnology tools can ass1st small farmers by developmg a range of technologies to add value and mcome to cassava farmers Severa! top1cs were d1scussed m the 40 plenary temal!c lectures and 125 posters presentations from wh1ch can be mentlon the followmg sc1enhfic h1ghhghts 1) Transgemcs are a reahty and are movmg rap1dly from the lab to the field 2) Genome mappzng With greater saturahon of markers neanng Idenl!ficatwn of CMV (cassava mosa1c virus) res1stance gene and extens1ve EST hbrar1es 3) Bzod¡verslty and populatwn genet1cs w1th vanat10ns m p1gments arch1tecture germmat10n cyanogens photosynthes¡s drought tolerance etc 4) ACMV (Afucan cassava mosa1c virus) w1th new vanants and satelhte genome 5) Post harvest detenoratwn funchonal genom1cs movmg forward 6) Double hapl01ds Idenhficatwn of recess1ve tralts to avmd genehc load 7) Expresswn profiles of pathogens zn cassava bactenal bhght 8) IPM and bwlogical control of msect pests 9) Improved mdustrwl vanetles m SE Asia and 1 O) In varo plants successful propagal!on of d1sease free plants for small farmer s groups m Colombia The plenary presentatwns posters and others CBN VI s documents can be found at CBN webs1te http 1/www c1at cg1ar org!bwtechnology/cbn/sixth_mtematJOnal_meetmg/mdex htm Participatwn m regwnal events to strengthen NARS capac1ty The CIA T Agrobwd¡vers¡ty and Bwtechnology ProJect had a Significan! partiCipatwn at the meetmg of Red810 held m Dom1mcan Repubhc on June 21 25 2004 A broad diversity of themahc areas were covered wh1ch ranged from stud1es on DNA sequence and gene express10n to apphcal!ons w1th small farmers m the field and mcluded the followmg top1cs structural and functlonal genom1cs positJOnal gene clorung marker ass1sted selectlon (MAS) 25 molecular plant/ pathogen mteracllons molecular breedmg for tolerance to ab10llc/ res1stance to b10l!c stresses plant genellc transformat10n m v1tro propagallon b10technology for classroom and small farmers bJOfortificatJOn and b10safety ProJect staff had 16 presentat10ns m symposmm or workshop and presented 16 posters Severa! proJect staff were part of the organ1zmg comm1ttee or chrurs of sess10ns The II Colomb1an Congress on B10technology was held m The Camara de Comercio and Uruvers1dad Nacional Bogota from 1 to 3 September 2004 CIA T s AgrobJOdivers~ty and 810technology ProJect partiCipated m the agncultural sess10n of the Congress g¡vmg 4 oral presentat10ns and presentmg 14 posters CIA T also participated m the Pre Congress event g¡vmg 13 lectures on Issues related to genom1cs and transgemc organ1srns The Pre Congress was open to the general pubhc and more than 200 students attended G Problems encountered and the•r solutwns Sorne of the maJar problerns facmg the proJect are similar to the encountered m prev10us years {space access to gene) whiie others surfaced dunng the year Salary of natiOoal recru1ted staff On the maJOr strength of the proJect 1s the quahty of the research ass1stants and assoc~ates mvolved However the leve! of salan es for severa! categones IS well below the average CIA T researchers As an exarnple based on analys1s made by human resources 7 out of the 9 research ass1stants 1 are below the CIA T average Discussions have been Imtlated between the staff of SB 2 and Human Resources to analyze the Situallon and Identify poss1ble solut10ns to be proposed to rnanagernent Mamtenance of maJor equ1pment The mamtenance of the maJar eqmprnents 1s becommg substandard due to lack of funds and lack of quahfied personal on stallon CIA T rnruntenance budget whiie well managed 1s too srnall cons1denng the nurnber of requests The lack of preventlve rnamtenance IS causmg delays for severa! proJects To ensure proper uses of the maJor eqmpment trammg sess10ns was Irnplernented for the newcomers DJscussJOns wllh sorne of the rnanor prov1ders are on gomg to tap mto therr expert1se for mamtenance Access to genes and freedom to operate Drscuss10ns wrth the pnvate sector have progressed to obtam access to elther mformallon or constructs However no maJar progress was rnade to obtam Bt constructs A consortrurn wrth Colombra was formed wrth the support of CIA T to explore the possibihlies of rdenllfymg useful Bt strams More lime and efforts need to be ded1cated for the negotlallon of possrble agreernents wrth the pnvate sector Space avallabrhty Lab and offices spaces contmue to be a rnaJOr constramt facmg the drfferent secllons ofthe proJect The avarlabrhty oflab and desk space for ass1stants students and vrsrllng screnllsts rs becommg hard to manage The purchase ofnew equrpment fundmg of new specral proJects and the mcorporallon of a new staff are the maJor causes of such constramt The restructunng made m 2002 has proveo to be msufficrent The proJect wrll contmue to seek effic1ent use of the lab space and as stated last year no obvrous solut10n seems m srght 26 Solut10ns to constramts razsed m 2002 2003 Space w1th the help of a spec1al emergency funds approved by the d1rector of research a the cassava genetlcs lab was remodeled prov1dmg a better quahty of the fac¡hlles and 1mproved use ofthe space However the team needs more space Cassava Frog Skm d1sease SB 2 has contmued to prov1de to the other sectwns of CIA T clean matenals from m v1tro cultures as the problem 1s restnctmg the ab1hty of the team to conduct field tnals w1thout the concems of delays and mfectwns Sorne progress was made m the area of d•agnostlcs by the relevan! proJect at CIAT workmg on the problem B1omformatJcs The 1ssue was ra1sed m 2002 Advances m the LIMS 1mplementat10n add¡llonal resources from the Generatlon Challenge program and trammg of local staff ha ve 1mproved the capac1ty of the group to tap mto the vanous databases In addJtlOn the team benefited from a VIS!! from IRD staff Benmt P1egu who set up for the team an effic1ent p1pehne for sequence analys1s In add1tlon the proJect 1mtlated w1thm Colombia a collaboratlve consortlum on bwmformatlcs wtth key mstltutwns such as Cemcana Cemcafe Cempalma Parque del Software and the Umv Nacwnal atmmg to share expertlse H Plans for next year Teams members wtll lead the tmplementatlon of the CIDA newly funded regtonal proJect on Combatmg Htdden Hunger m Latm Amenca Btoforttfied Crops wtth Improved V¡tamm A Essentlal Mmerals and Quahty Pro te m The collaboratlve ts a partnershtp between CIA T CIP C1mmyt Embrapa and Clayuca and wtll mvolve NARS from Central Amenca and the Andean regwn Team members wtll contmue the1r actlvtltes wtthm the HarvestPlus challenge program m the area of 1ron and zmc btoforttfed bean and beta caroten cassava Teams members w!ll also mteract wtth the Generatlon challenge program and parttctpate m the 1mplemetatlon of the dtfferent clusters estabhshed m the CP etther thru commtssed research or competttve grants In the area of transgemc research the team members wtll contmue to develop the tools for nsk assessment and gene flow studtes for nce and bean as part of the btosafety nsk assessment proJeCt The tssue of deregulatton of transgemc wtll be analyzed and a strategy wtll be formulated to move CIA T product towards field and food safety testmg In the area of new tools mtcroarray technolog¡es already tmplemented and Smgle Nucleottde Polymorphtsm (SNP) Wiil contmue to be tncorporated mto the screerung of germplasm and marker ass1sted selectton Mtcroarray and real time PCR analys1s wtll be expanded to gene expresston studtes to better understand abtotlc and btotlc stresses V IIliS Induced Gene Stlencmg (VIGs) protocols Will be tmplemented to asstst m the gene clonmg of dtsease reststance genes The use of the untgene set of cassava wtll be expanded Addtltonal mtcroarray chtp for abtoltc stress for nce and beans and post harvest detenoratton m cassava wtll be developed 27 The unplementat10n fo the project strategy to mtegrate CIAT work on biOtechnology m Afnca With breedmg will be expanded BIOtechnology proJects w1th colleagues m Rwanda are bemg developed m the area of MAS m beans and Cassava The work already mt1ated With RF fundmg on cassava MAS m Tanzan1a Will be expanded thru additiOnal trammg and SI te VISits A newly funded proJect on biOtechology trammg m Southem Afnca from USAID to CIAT and M1clugan S tate Umvers1ty will prov1de the means to Im!Iate the collaboratiOn Afncan NARS Collaborations w1th NARS m Latm Amenca Wlll be pursued m the area of genellcs genom1cs !Issue culture and genet1c transformatiOn Pubhc awareness m the area of biOtechnology and biOsafety for pohcy markers and journahsts will contmue 1 ProJect Performance lndicators 1 TECHNOLOGIES METHODS & TOOLS 1 1 Genes tagged more than 12 genes and QTLs Identified m bean nce cassava and Brachtarza 1 2 Methods tmplemented M1croarray fulllenght cDNA hbranes SubstractiOn cDNA hbranes constructlon Smgle Nucleotlde Polymorphism real time PCR BAC sequencmg and annotatiOns B10safety field assessement sequence annotatlons and analys1s DArT 1 3 Support Tools Ltbranes developed Gene Ltbranes/Construct cDNA A root spec1fic cDNA hbrary was constructed from a substractlve hybndiZatiOn usmg cDNAs from the aenal part (Leaves & stem) as dnver A m1x of five wh1te rooted culhvars were selected to make the hbrary 768 clones form potato leaves varelles 703508 CHS 625 PS 3 704746 705428 700347 (CIP number Idenllficatlon) DNA d1gested wtth Pstl and Taql for evaluatmg d1verstty w1th DArT 5760 clones from potato leaves varetles 703508 CHS 625 PS 3 704746 705428 700347 (CIP number IdentlficatiOn) DNA d1gested wilh Pstl and Apol for evaluatmg divefSity w1th DArT 28 Cassava Uo¡geoe Set Two economically Importan! characters starch content and Cassava Bactenal Bhgth res1stance were targeted to generate a large collectlon of ESTs Two hbranes were constructed from cassava root tlssues of van elles w1th h1gh and low starch contents (MPER 183 and CM523 7) the number of generated sequences of each hbrary was 5 376 and 4 992 respectlvely that resulted m 4 992 umque sequences Other hbranes were constructed from plant tlssues challenged by the pathogen Xam and were generated 4134 umque sequences In total a cassava umgene set w1th 5 700 sequences was constructed lb ry C\1 MP \1C 1-48h MB mb_ o b mb_ds sg_rw b g_ssh sg_dsc: m sh O'IA AFLP Ge b k Total DNA No q g t d 5,)76 4992 2,304 2 688 384 768 288 384 768 384 18 316 N e es S gl to (/) Te t t A lyz d 3 608 555(15) 3 391 1 127 (33) 1 721 1 178 (68) 1 560 1 049 (67) 258 124 (48) 438 41 (9) 128 95 (74) 210 128 (61) 382 17 (5) 2S8 142 (SS) 241 179 (74) 848 534 (63) 13 043 3 825 BAC Two BAC hbrmres for bean and cassava Data Bases umted/1mproved e t gTC u g (/) 642 1197(33) 607 1 734 (51) 184 1 362 (79) 158 1 207 (77) 22 146 (57) 54 95 (22) 6 101 (79) 24 152 (72) 29 46 (12) 45 187 (73) 27 206 (86) 98 632 (75) 1 875 5 700 A Database from cassava with a total of 11 954 cDNA sequences from the 5 ends and 111 from the 3 ends All sequences were clustered m 5 700 umque sequences Currently th1s database 1s avatlable locally at CIA T and the user can perforrn sequences retneval and batch BLAST search Pubhcatwos RefereedJoumals Pubhshed 22 Refereed JOUmals submttted 21 Book chapters Pubhshed 6 29 Strengthenmg NARS The proJect concentrated its efforts this year on strengthenmg NARS through a senes events either at CIA T headquarters or at NARS facliitles A total of more than 562 persons from natwnal and mtematwnal mstltutwns recelVed trammg with SB 2 ProJect Staff or partiCipated m SB 2 led events 1 Sixth mtematwnal Scientlfic Meetmg of the Cassava Bwtechnology Network March 8 14 2004 ( 150 part1c1pan ts) 2 Introductlon to bwtechnology for Syngenta - Colombia Bwtecnolog1a Bas1ca y Plantas Modificadas Genetlcamente I Taller Syngenta March 26 2004 (21 parttc1pants) 3 Trammg on m vltro tlssue culture to farmers assoc1at1ons Nuevas vanedades de yuca maneJo y control de la mosca blanca y el cuero de sapo Caseta Comunal de la Vereda Alegnas Santander de Qmlichao Cauca August 25 2004 (70 partlcipants) 4 Introductlon to genom1cs and transformatwn one Day Pre Congress Workshop for bwlogy students on Genom1cs and Transformat10n Bogota Umvers1dad Nacwnal August 31 2004 (200 part1c1pants) S Bwmformat1cs May 28 04 (15 part1c1pants from CeDJcana Ce01cafe CeDJpalma Parque Software) 6 Planmng workshop on Combatmg H1dden Hunger m Latm Amenca Bwforhfied Crops w1th Improved Y1tamm A Essentlal Mmerals and Quality Protem Sept 29 Oct 1 04 ((51 part1c1pants) 7 Bwsafety workshop for natlonal organ¡zatlons and mmistnes m Colombia Oct 4 6 2004 (43 part1c1pants) 8 Gene flow and bwsafety workshop Oct 7 9 2004 (41 part1c1pants) 9 Workshop on bwtechnology and genetlc transformat10n for Sciences and Agnculture commlttee ofthe Colombian Congress Oct 7 9 2004 (12 part1c1pants mcludmg s1x senators from tbe Colomb1an congress) Strengthenmg tbe tecbDJcal capacJtles of CIA T SB 2 ass1stants In additlon to strengthemng the capaclty of NARS the proJect put emphasiS this year on trammg CIAT SB 2 assistants The mechanism for the trrumngs was either thru site ViSits to CIAT collaborators m the US and France to leam new techniques or thru attendance of selected courses It iS worth mentwmng that as of 2005 5 CIA T assistants managed to get accepted With either full or partial fellowship to attend severa! of the very compehtlve courses provided by the Cold Spnng Harbor lab 1 Mauncw Soto full fellowships to the Workshop on NSF Rice Oligonucleotlde Array The Instltute for Genomic Research (TIGR) Rockvlile MD September 20 22 2004 2 Catalina Romero part1al fellowship to the Advanced Techruques m Plant Science Cold Spnng Harbor USA July 1 21 2004 3 Juliana Chacon partial fellowship to attend the Workshop on Molecular Evoluhon Woods Hole Mass July 26 Aug 6 2004 10 28 30 4 Paul Chavamaga Trammg m Richard Sayre s Lab at Oh10 State Umverstty Follow up on USAID funded proJect Expandmg the range of uses of cassava starch A source ofmcome generatwn OH USA February 2004 5 Mynam C Duque Summer Instttute m Stattsttcal Genettcs at North Carohna State USA Introductton to Genomtc Sctence May 26 y 28 04 Introductton to Bwmformattcs June 2 4 04 Mtcroarray Analysts June 7 9 04 6 Roosevelt Escobar Vtstt to Richard Lt!z Laboratory on ttssue culture for frmts U Flonda USA Nov 17 2003 7 Ehana Gattan btomformattcs trammg at Comell Umverstty USA July 2004 SNP trammg at the USDA soybean lab Dr Perry Cregan Belstvtlle March 2004 8 Fernando RoJaS Parttctpatwn m the Consultatton Workshop for Subprogram 4 mformattcs m the Challenge Program unlockmg Genettc Dtverstty m Crops for the Resource Poor IPGRI - Rome Parttctpatton m the Workshop for Subprogram 4 - Generatton Challenge Program Informatton Systems Platform & Network Destgn CIMMYT - Mextco Parttctpatwn m the Generatton Challenge Program (GCP) Informatwn Systems Platform and Network lmplementat10n Workshop IRRI - Phtlhpmes 9 Morgan Echeverry Trammg m Real Ttme PCR Btomol Laboratory Bogota Apnl 2004 3 2 lndtvJdual trammg 31 NARS researcher rece1ved trammg w1th SB 2 staff (Oct /03- Oct 04) 3 3 V1S1tors Aprox 430 persons vtstted the facthtles of the proJect whtch amount to 43% the total of CIA T vtsttors 3 4 Current graduate students PhD 6 MSc 5 Undergraduate 18 Completed Thests 14 J Resource Mob•hzabon ProJect approved or on gomg • Htgh through put genettc dtverstty charactenzatton of germplasm wt!h a DNA chtp Donor IPGRI 31 • Development of strateg1es for better targetmg of seed rehef and hnkmg rehef and rehabihtatwn Donor FAO • Bwfortdied Crops for Improved Human Nutnhon Donor WB/IFPRI • Bean Genom1cs for Improved drought Tolerance m Afnca and Latm Amenca Donor GTZ • Genetlc Mappmg of the Lmamann bwsynthet1c genes CYPDI and D2 and the development of markers for CNP m Cassava m collaboratlon w1th Prof B1rger Moller Royal Agnculture and Vetennary Umvers1ty Copenhagen (DANIDA) • Genoplante ProJect for phenotyp1c and charactenzatwn of a senes of T DNA mutants Evaluatlon and multiphcatwn of 5000 !mes de T DNA mutants • An mtegrated approach for genetlc Improvement of alummmm resistanse of crops on low fert1hty ac1d sOI!s • Knowledge and tools for the modulatwn of post harvest physwlog¡cal detenoratwn m cassava • Seed a1d and germplasm restoratwn m d1saster Situations Smtes1s of Lessons leamed and promotwn of more effective practices • Model of food safety assessment of transgemc crops • Bean genom1cs for Improved drought tolerance m Afnca and Latm Amenca • Dehvery of transgemc nce cultivars to seed producers and farmers m Tropical Amen ca • Followmg a multi step approach mvolvmg bwsafety assessment mutntwnal testmg and negohatwns • Gmes Mera memonal fellowslup fund for prostgraduate stud1es m bwdiversity • Cassava bwtechnology network IV • Gene flow anahs1s for assessmg the safety of810 eng¡neered crops m the tropics • Combatmg h1dden hunger m Latm Amenca BIOfort¡fied crops w¡th 1mproved V1tamm A essential mmerals and quahty protem A collaboratiVe proJect subm1tted to CIDA on behalf of A Partnerslup of InternatiOnal Agncultural Research Centers and NatiOnal Agncultural Research Systems m Latm Amenca • Development ofm1cro satelhte markers to fac!lltate use ofthe cassava 32 • Molecular genetlc maps by Afncan collaborators workmg on gene mappmg resistanse toCMD • Towards the development of mdustnal cassava vanetles genetlc and molecular anahsis of early bulkmg m cassava germplasm collectlon • Sustamable ml palm product10n as a source of employment and mcome for rural commurutles and small scale farmers m tropical Latm Amenca • Molecular CharactenzatiOn of Genetlc Diversity and the Defimt10n of Heterotlc Groups m Cassava • Developmg and explmtmg expressed sequense Tags for cassava Starch and Bactenal Bhght Resistanse • Expandmg the range of uses of cassava starch A so urce of m come generat10n • Seed aid and germplasm restoratlon m disaster SituatiOns smtesis of lessons leamed and promot10n of more effective practices • Development of an m VItro protocol for the product10n of cassava doubled haplmds and Its use m breedmg • Comparatlve genomics and genetics m legumes a collaboratlve research ProJect between CIA T and Umversity of Aarhus concept note prepared for DANIDA • Phaseomics wRUIG GIAN (Submltted by Umv Of Geneva with CIAT collaboratlon) • UtihzaciOn de luerro y zmc en modelo animal y respuesta chmca al consumo habitual de fnjol de alta densidad romeral en muJeres y mnos Submitted by UruvefS!dad del Valle (with CIAT) to Colciencias • The molecular diVefS!ty network ofCassava (MOLCAS) • Mutagenesis of Cassava (Manihot esculenta Crantz) for the generatlon IdentificatiOn and Molecular Anahsis of Novel trmts Research Contract submitted to the Intemat10nal Atomic Energy Agency (IAEA) VIena Austna • Challenge Program Unlockmg Genetic Resources m Crops fort the Resource Poor • Development of an In vitro Protocol for the Product10n of Cassava Doubled Haplmds and Its Use m Breedmg Submitted to ZIL Swltzerland 33 Proposal and concept notes • Rice Improvement for the Canbbean Reg10n through molecular breedmg • Understandmg genetlc diVersity of cassava (Manihot esculenta) and wild relatives for broademng the crops genetlc base and sustamable agncultural development • Product10n of Quahty plantmg matenal of tropical frults through b10technology to supplement small farrners mcome m the Colombian Llanos • Producc10n entable y sostemble de guanabana mediante el desarrollo de tecnolog~as de post cosecha y el aprovechamiento de subproductos • Combatmg hidden hunger m Latm Amenca 810fortlfied crops With Improved vitamm A essentlal mmerals and quahty protem • Gene discovery for Alummum resistance m common bean and Brachiana • Light water and stomata gene targets for multlple ab10t1c stress tolerance • Development of a common plant stress DNA chip and Its apphcat10n for crop Imrpovment • Request to Syngenta for Bt genes or other msectiCidal genes available to produce transgemc cassava plants that help control the homworm Ennn)'ls ello m Colombia Prepared by Paul Chavamaga and Manabu Ishitani February 2004 • Flowers Fruits and Roots Modlficat10n of Flowenng to Improve Trruts of Agncultural lmportance A Proposal Submltted to the Rockefeller Foundat10n Prepared by The Department of Plant Developmental B10logy Max Planck Instltute for Plant Breedmg Research Cologne 50829 and The Intemat10nal Center for Troptcal Agnculture AA 6713 Cah Colombia Apnl2004 • Engmeenng the Plastid Genome of Staple Crops for High Express10n and Contamment of Transgenes A proposal submitted to the Generatlon Challenge Program • Improvement of lron Zmc and Protem Levels m Crops Usmg Selected Transgenes A proposal submitted to the Global Challenges m Global Health Imtlatlve of the Nat10nal Instltute of Health • Engmeered chromosomes for dehvenng sets of genes to staple crops Submitted to the Grand Challenge m Global Health lnitlatlve ofthe Nat10nal mstitute ofHealth • Cooperatlve Project to Enhance and Exchange Technolog¡es for Producmg High Quahty Seed of Staple Crops m Uganda Kenya Rwanda and Colombia A concept note submrnited to the Rockefeller Foundatlon 34 • InterdiSCiplmary Center for Non Conventwnal Breedmg of Crops Relevant to Colombia (ICNCB) A proposal submitted to COLCIENCIAS seekmg fundmg for the creatwn of a Center of Excellence m Colombia 35