Plant Breedingand Genetics

Newsletter

______________________________________________________________________Joint FAO/IAEA Division of Nuclear Techniques in Food and Agricultureand FAO/IAEA Agriculture and Biotechnology Laboratory, SeibersdorfInternational Atomic Energy Agency, Viennahttp://www.iaea.org/programmes/rifa/http://www.fao.org/WAICENT/Agricul.htm

No. 1June 1998

CONTENTS

TO THE READER........................................................................................................................1

A. STAFF ........................................................................................................................................2

B. FORTHCOMING EVENTS ...................................................................................................3

C. PAST EVENTS .........................................................................................................................3

D. STATUS OF EXISTING .......................................................................................................11

E. NEW CO-ORDINATED RESEARCH PROJECTS .........................................................14

F. ACTIVITIES AT THE PLANT BREEDING UNIT, SEIBERSDORF...........................21

G. PUBLICATIONS ...................................................................................................................23

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TO THE READER

Dear Reader,This is the first issue of the Plant Breeding and Genetics Newsletter. The Newsletter will

inform you about current activities of the FAO/IAEA sub-programme on plant breeding andgenetics which is implemented by the Plant Breeding and Genetics Section of the JointFAO/IAEA Division of Nuclear Techniques in Food and Agriculture (Vienna) in closecollaboration with the Plant Breeding Unit of the FAO/IAEA Agriculture and BiotechnologyLaboratory (Seibersdorf). The work of this sub-programme goes back to 1964 when, with theestablishment of the Joint FAO/IAEA Division, activities were initiated to realise the potentialof different types of ionizing radiation as well as radiomimetic compounds to induceagronomically useful mutations in crop plants.

Interest in the use of mutation techniques to generate and select desired genetic variationin crop species has increased significantly during the last decade. This has been mainly dueboth to the substantial success in applying in vivo mutation techniques in breeding new,improved varieties and to the new opportunities for induced mutations in vegetativelypropagated crops using in vitro techniques and the advances made in rapid, often non-destructive, mass-screening methods. Also, recent developments in plant molecular geneticshave indicated that induced mutations are probably the most effective approach to generatedesired genetic polymorphism in plant characters and hence mutational analysis of plantstructure and function has become the main interest of many laboratories. On the basis of thesedevelopments the Section now implements activities within the framework of the followinggeneral mandate: To support national plant breeding programmes and enhance biodiversitythrough applying mutation techniques and modern biotechnologies to improve local, oftenneglected major food crops in marginal and stress-prone areas, and to domesticate plant specieswith potential value for food or export products in partnership with National AgricultureResearch Systems (NARS) and International Research Centres.

Already in the early 70's the need was expressed by numerous co-operators forcirculation of a Newsletter on induced mutations and their applications in plant breeding andgenetics, and the Section followed this up by publishing the first issue of its Mutation BreedingNewsletter (MBNL) in May 1972. Over the following 20 years, the MBNL was edited byAlexander Micke, the then Section Head (1969-1991). Over the years the MBNL, developedinto a scientific journal and due to increasing number of scientific papers, essentially no spacewas left to provide researchers with information about the activities of the sub-Programme onplant breeding and genetics. This information gap has become increasingly apparent as ouractivities have expanded into the application of in vitro, molecular methods and other relatedbiotechnologies in Co-ordinated Research Projects, Training Courses or Technical Co-operationprojects. To fill the information gap we have therefore decided to publish this Plant Breedingand Genetics Newsletter (PBGN) every 6 months so that you have more information aboutcurrent and planned activities of the FAO/IAEA Plant Breeding and Genetics sub-Programme. The newsletter is also available on the Joint Division's home pages on the Internet. I would liketo emphasize, however, that this Newsletter does not replace the MBNL. In fact, the MBNLwill continue to publish scientific papers related to the application of mutation techniques inplant breeding and genetics.

Miroslaw Maluszynski

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A. STAFF

Joint FAO/IAEA Division of Nuclear Techniques in Food and Agriculture,Vienna International Centre, P.O. Box 100, A-1400 Vienna, Austria.

Jame s Dargie Director E-mail : J.Dargie@iaea.org

Manase Pete r Salema Deputy Director E-mail : M.P.Salema@iaea.org

Plant Breeding and Genetics Section

Miroslaw Maluszynsk i Head of Section E-mail : M.Maluszynski@iaea.org

Beant Ahloowalia Technical Officer E-mail : B.Ahloowalia@iaea.org

Karin Nichterlein Technical Officer E-mail : K.Nichterlein@iaea.org

Leo van Zanten Technical Officer E-mail : L.Van-Zanten@iaea.org

Amram Ashri Technical Officer E-mail : A.Ashri@iaea.org

Kathlee n Weindl Secretary E-mail : K.Weindl@iaea.org

Katayo n Entekhabi Secretary E-mail : K.Entekhabi@iaea.org

FAO/IAEA Agriculture and Biotechnology Laboratory, A-2444 Seibersdorf,Austria

Christophe r Rigney Acting Head E-mail : C.Rigney@iaea.org

Plant Breeding Unit,

Javier Zapata Aria s Head of Unit E-mail : F.Zapata-Arias@iaea.org

Paolo Donini Researcher E-mail : P.Donini@iaea.org

Nicolas Roux Researcher E-mail : N.Roux@iaea.org

Marcella Guzman Junior Professional Officer E-mail : M.Guzman@iaea.org

Stefan Nielen Associate ProfessionalOfficer

E-mail : S.Nielen@iaea.org

Rownak Afza Researcher E-mail : R.Afza@iaea.org

Andrea Kodym Technician E-mail : A.Kodym@iaea.org

G. Berthold Technician

A. Draganitsch Technician

F. Zwiletitsch Technician E-mail : F.Zwiletitsch@iaea.org

M. Weinreich Secretary E-mail : M.Weinreich@iaea.org

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B. FORTHCOMING EVENTS

Third and final ResearchCo-ordination meeting on" In vitro Techniques forSelection of RadiationInduced Mutants Adaptedto Adverse Environmen-tal Conditions".17-21 August 1998,Shanghai, China.

Third and final ResearchCo-ordination meeting on"Induced Mutations andOther Advanced Tech-nology for Production ofCrop Mutants Suitable forEnvironmentallySustainable Agriculture".24-28 August 1998,Stuttgart, Arkansas, USA.

Third and final ResearchCo-ordination meeting on"Induced Mutations inConnection with Bio-technology for CropImprovement in LatinAmerica". 5-9 October1998, Lima, Peru.

C. PAST EVENTS

Final RCM of the FAO/IAEA/Italy Co-ordinated Research Project on Improvement ofBasic Food Crops in Africa through Plant Breeding, Including the Use of InducedMutations. Naples, Italy, October 30-November 3, 1995.

This Co-ordinated Research Project,funded by the Italian Government, wasinitiated in 1989. The primary objective wasto breed improved varieties of staple foodcrops of Africa with emphasis on theindigenous species and their local cultivars.Twenty persons participated from BurkinaFaso, Cameroon, Côte d'Ivoire, Ethiopia,Ghana, Kenya, Liberia, Mali, Nigeria,Tanzania, Uganda, and 14 scientific reportswere presented.These included reports from 10 ResearchContract holders from Africa, 3 TechnicalContract holders from Italy and an updateon research carried out at the FAO/IAEAAgriculture and Biotechnology Laboratory,Seibersdorf. The proceedings of the meetingare available as IAEA TECDOC-951 andcan be obtained free of charge by writing tothe Section. We hope that this documentwill be of value to researchers, students andpolicy makers alike in their endeavor topromote plant breeding and increase foodproduction in Africa.

Workshops Under a regional Technical Co-

operation project (RAF/5/035) on thecontrol of Bayoud disease of date palm,three workshops were held. The first was on"Mutation induction and selection by usingin vitro techniques", 6-17 November 1995at FAO/IAEA Agriculture andBiotechnology Laboratory, Seibersdorf.The second was on "Development of invitro techniques for mutant selection indate palm", and was held at INRAT,Degache, Tunisia, 20-25 May 1996. Sevenresearchers from Tunisia, Algeria andMorocco participated. The third workshopon "Use of molecular techniques for mutantselection", was held at INRA, Marrakech,Morocco, 10-21 March 1997. Sixparticipants from Tunisia, Algeria andMorocco participated and it includedlectures and laboratory exercises on DNAisolation, restriction, amplification, PCRand RAPD techniques.

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Consultants Meeting on Bayoud Disease of Date Palm, IAEA, Vienna, 25-29 March 1996.

Date palm plays an important role in the economic and social life in the Sahara. There isan old saying that "without date palm there is no life in the Sahara". Bayoud disease of datepalm is caused by Fusarium oxysporum f.sp. albedinis (F.o.a). This disease has alreadydestroyed 15 million date palm trees in North Africa and has become a trans-national problemrequiring a trans-national solution, involving a free exchange of information and experiencebetween participating countries and international technology transfer.

This consultants meeting was held to discuss Bayoud disease in depth and makerecommendations to control it. The group considered current knowledge on the fungusincluding: biotypes; the spread in North Africa; methods of early detection; rapid diagnostictechniques in mature palms; tests for resistance based on pathological, genetic and molecularmethods; fungal toxin characterization to differentiate between biotypes and for co-cultureexperiments. The group made recommendations on the short and long term strategies to breedfor resistance to Bayoud, including induction of variation for resistance to Bayoud in wellestablished date palm varieties using induced mutation and in vitro techniques.Recommendations were made to develop procedures to select for resistance to Bayoud based ontoxin co-culture in vitro. A document based on the recommendations was published and can beobtained by writing to the Section.

Regional Workshop on Evaluation of Promising Cereal Mutants. INTA, Argentina, 21-25October 1996.

This workshop aimed at trainingscientists from national institutions onplanning and conducting multi-locationtrials of promising cereal mutants. Itcovered the principles of designing andorganizing regional multi-location trials,seed multiplication, exchange of mutantgermplasm, collection of data related toagronomic performance of mutant lines,statistical evaluation of the data andanalysis. The workshop programmeconsisted of lectures and round tablediscussions. The participants presentedreports on the implementation of the

programme agreed during an earlierworkshop held in Campinas, Brazil, from 4-8 December 1995. The workshop provideda forum for discussing problems related toexchange of germplasm among theparticipating countries. Biological tools forshortening breeding cycles were alsodiscussed. The participants also presentedtheir work plans for the next year of theproject which were then discussed toestablish a uniform design of the trials andprocedure for evaluation of mutants. Areport on this workshop is in preparation.

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Final RCM of the FAO/IAEA Co-ordinated Research Project on The Application of DNABased Marker Mutations for Improvement of Cereals and Other Sexually ReproducedCrop Species, Vienna, Austria, 4-8 November 1996.

In recent years, great strides havebeen made in the development of molecularmarkers suitable for studies of sexuallypropagated crop species. Markers that arebased on the polymerase chain reaction(PCR) technique are suitably polymorphicfor applications in breeding programs andwill be used increasingly in crops importantin both developing and developed countries.

The most commonly used DNAmarkers are restriction fragment lengthpolymorphisms (RFLPs) and randomamplified polymorphic DNAs (RAPDs). Another marker technology, known assimple sequence repeats (SSRs) or "micro-satellites”, has many desirable features,including the use of a PCR reaction insteadof a blotting procedure, co-dominantinheritance, and genome-specificity inpolyploids, but is expensive and time-consuming to develop due to therequirement for DNA sequencing. Yetanother new marker technique has recentlybecome popular, namely AmplifiedFragment Length Polymorphisms (AFLP). The AFLP technique is based on theselective PCR amplification of restrictionfragments from a total digest of genomicDNA. Finally, the newest type of DNAmarker is known as the "biochip". Theseare high-density oligonucleotide arraysbound to a solid microchip. The method canbe used to characterize the spectrum ofsequence variation in a population and canbe applied to the analysis of many genes inparallel. Since biochips are still highlyexperimental and expensive to design, itmay be some time before they arewidespread in crop genetic systems.

The conclusions and recommendationsfrom this project are the following:- Different types of molecular markers are

best suited to different types of mappingand breeding applications. Choosing themost appropriate marker system is essentialfor success. Effective planning andexecution in the capture and management ofmarker data is also critical.- Genetic markers, such as phenotypically-identified macro-mutants (those which havea large effect on the phenotype) andcytological markers can provideopportunities for scientists to assess theagricultural significance of a gene orcharacter. Integrating this information withthat derived from molecular markers makesmacro-mutants even more significant forpractical purposes, as shown for tomato andpotato. Integration of cytologicalinformation with DNA markers enhancesthe understanding of genome architecture.- The identification of mutations withlinkage to RFLP, PCR or DNAfingerprinting markers has further increasedthe practicability of using mutants asexemplified in downy mildew resistancegenes in lettuce where the relation betweendeletion breakpoints and molecular markershas provided greater resolution than meioticrecombination.- Integrated mapping resources - includingcomparative maps between related cropspecies, genome databases, and clone banks- have been created. These resourcesenhance the utility of molecular markers inplant breeding and mutation analysis.- In the long-run, the most powerful use ofmolecular markers in plant breeding may bethe ability to clone genes known only byphenotype. It must be realized, however,that the cloning of genes of agronomicinterest is currently an expensive,demanding enterprise which is onlypossible in the most advanced and wellfunded laboratories.

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- Types of cloning strategies such astransposon tagging that directly mark sites,rely on efficient transposon activity in thespecies of interest and have worked well inmaize and tomato, however this may not begenerally applicable. Agrobacteriummediated transformation is being utilized ina number of crop species, but itsapplicability is limited to crops whereefficient transformation and regenerationand efficient means of screening for thedesired mutants are available. - The end result of identification of genesof economic interest is their application inthe production of commercial materials. Transformation utilizing genes of economicinterest is now a reality for many crops. Apotential use of physical mutagenesis is toremove unwanted selective markers in thelater stages of transgenic line evaluations.- The primary obstacles to widespread useof markers are the lack of facile markersystems and the resources required tosupport their use. The advantages providedby markers are many but they depend onnumerous variables such as crop biology,trait(s) undergoing selection, resources, andtype of cultivar. However, it has becomeclear that markers should be considered asan option under many circumstances andaspects of crop improvement. - Monoclonal and polyclonal antibodieshave been routinely used for strainidentification with viruses. It helps theassessment of the homogeneity and stabilityof the pathogen population, introduction ofnew strains and purity of inoculum. However, there are limitations to extendingthese techniques to other pathogens and thedevelopment of PCR-based DNA markerswould make important contributions to cropimprovement by clarifying the geneticarchitecture and repertoire of the pathogenpopulations.- The majority of agriculturally importantcharacters are multigenic, stronglyinfluenced by environment and expensive to

evaluate directly. Enormous progress hasbeen made by DNA marker techniques, butthe inability to describe and select forspecific quantitative trait loci (QTL) maylimit the breeders’ ability to make progressin the future.- Given the rather recent development of thesuite of enabling technologies, adoption andimplementation of molecular techniques hasbeen very rapid. Many other cultivars willbe developed through these methods fortraits such as grain quality (starchproperties, oil quality, protein quality),disease resistance, insect resistance,exploitation of heterosis (throughengineered nuclear male sterility). Thesemethods not only facilitate the productionof novel cultivars, but also the production ofcultivars in far less time.- However, many challenges remain forcomplex (multigenic) characters such asdrought-, cold- and salt tolerance, and yield.Considerable progress has been made usingDNA markers to tag agronomicallyvaluable genes in many crop species andthus the foundation has been established forusing those markers and mutations for morerapid cultivar development in specieslargely ignored by the private sector.Virtually all phases of plant breeding,including selection of parents, prediction ofprogeny performance, progeny selectionand varietal identification, have beenaffected by these tools. The prospects arevery good for markers to positively affectthe rate of genetic gain. The criticalcomponent then, will be maintenance andcreation of genetic variation through thevarious methods.

The final technical document of thisCRP was published at the end of 1997.

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FAO/IAEA Regional Training Course on Molecular Approaches, Mutations and OtherBiotechnologies for the Improvement of Vegetatively Propagated Plants, Kuala Lumpur,Malaysia, 28 October - 8 November 1996.

15 trainees from China, India,Indonesia, Malaysia, Pakistan, Philippines,Republic of Korea, Sri Lanka, Thailand andVietnam participated in this course whichcovered the use of chemical and physicalmutagens and their mode of action; theinduction of mutations and selection underin vivo and in vitro conditions; micro-propagation methods for separation ofchimeras and rapid multiplication; and the

use of molecular methods and DNAmarkers in mutant identification andselection. These topics were handled by thelecturers Dr. O. Kamra (Canada), Dr. R.Litz (USA), Dr. G. Kahl (Germany), Dr.Mohammed Bin Osman, Dr. Mak Chai, Dr.Normah, Dr. Nazir and Dr. Farida Shah(Malaysia). We thank the lecturers and thecourse director Dr. Zakri for their excellentcontributions to the success of this course.

Consultants Meeting on Biotechnology in the Next Millennium, Vienna, Austria on 2-6December 1996.

Consultants from Colombia,Germany, Pakistan, South-Africa andrepresentatives from FAO and UNIDOexchanged visions and discussed the pastlessons and future strategies for rapidintegration of biotechnology in cropimprovement for the developing countries. During the meeting an overview was givenon the research strategy and programmeactivities of the Plant Breeding andGenetics Section of the Joint FAO/IAEADivision together with a broad overview onpast developments on biotechnologies in theAfrican, Asian and Latin American regions.

The recommendations from thismeeting can be used as guidelines for

biotechnology activities in developingcountries. These concern the creation ofawareness on national and regional levels,the commitment of national institutions tobiotechnology programmes, priority setting,the optimal utilization of already existingcapacities, capacity building, andnetworking at the regional and internationallevel. More specific guidelines wereformulated on technologies such as in vitroand molecular techniques, and thedevelopment of human resources. Theaspect of Intellectual Property Rights andthe issue of biosafety was included inrelation to the development of transgenics.

First Co-ordination Meeting of the Regional Technical Co-operation Project onDevelopment and Promotion of Improved Mutant Crop Varieties. Biotechnology andNuclear Agriculture Research Institute, Ghana Atomic Energy Commission, LegonAccra, Ghana, 3-7 February, 1997.

Project co-ordinators from Algeria, Cameroon, Côte d'Ivoire, Egypt, Ethiopia, Ghana, Kenya,Libya, Madagascar, Mauritius, Morocco, Sudan, South Africa, Sierra Leone, Tanzania, andZaire participated in the meeting and planned the national and regional activities under theproject, especially the evaluation of mutant lines/clones developed under previous technical co-operation projects, research contracts or national programmes. It was agreed that the following

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mutant lines/clones will be evaluated in 1997/98: sesame and safflower in Egypt, cotton andbanana in Sudan and cocoa in Ghana. Work plans were prepared and discussed.

Final RCM of the FAO/IAEA Co-ordinated Research Project on Use of Novel DNAFingerprinting Techniques for the Detection and Characterization of Genetic Variation inVegetatively Propagated Crops. Bhabha Atomic Research Centre, Mumbai, India, 24-28February 1997.

During this CRP, experience wasgained in building productive collaborationsbetween research scientists and breedersusing biotechnology as the commonlanguage allowing the breeder to explain hisproblems and the research scientist topropose strategies to resolve them. As aspin-off, the breeders and research scientistshave been endowed with capacities totransfer the knowledge and know-how. Themonologue between “Biodiversity Centers”and “Biotechnology Centers” has mergedinto a profitable dialogue. The pressure onmodern breeders caused by the urgentdemand for more and better diseaseresistant and high yielding food crops,increases the need for continuousimprovement of plant varieties. The basisof genetic improvement is the optimizationof gene interactions and is based on geneticdiversity, which can be obtained fromseveral sources, including: naturalpopulations; products of sexual crosses (viarecombination, segregation and selection);spontaneous mutations (aneuploidy,polyploidy, other mutations); inducedmutations (physical and chemical); and insertional mutagenesis (transposableelements, T-DNA, retroposons).

DNA markers are integrallyconnected with the success of molecularbreeding. Moreover, juvenile markers, i.e.markers evaluated at early stages of plantdevelopment, are needed by breeders indeveloping countries to identify new genesources in the available biodiversity, toselect parents in order to increase heterosis,to decrease the number of backcross

generations, for gene introgression breedingprogrammes, for marker assisted selection(MAS) and, ultimately, for gene isolationand transfer via map-based or deletion-based cloning. Fortunately, sufficientnumbers of powerful genetic markers havebeen developed in human biology.

PCR technology has boosted thescientific output and yield of this worldwidecooperative research. Based on PCRtechnology, neither a complex laboratoryinfrastructure nor highly trained and skilledstaff are required to effectively genotypeentire breeding populations with largenumbers of individuals, as proven bymarker assisted cattle breeding. PCRanalysis is very simple, robust and reliable:a minimal quantity of total genomic DNA,even partially degraded, is enough to act asthe template for multiplex amplifications. Germplasm assessment and genotyping foridentification purposes (breeder’s rights,variant authentication, conformity test) canbe achieved with polymorphic loci (geneticfingerprinting). The homogeneity of invitro mass propagated material can easily bemonitored using recurrent checks during thewhole process. PCR assays may be readilyautomated with no further humaninterference between sampling in situ (inthe fields) and analysis “in silico” (on thecomputer screen), which will allow thetransfer of molecular markers to the “grassroots”.

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Achievements of this CRP:Various molecular marker technologies were used to recognize genetic diversities in

vegetatively propagated crops. The building of this research network permitted the transfer oftechnology and the enrichment of human and genetic resources available to individual projectswhere technologies of DNA profiling (RFLP, RAPD, SSR, AFLP, DAF, and RAMPO) wereevaluated. Cost, convenience, reliability and information content were recognized as key criteriafor selecting an appropriate profiling technology.

The association of markers and morphological traits led to the generation of maps inseveral species which allow the exploitation of alternative life cycles of vegetatively propagatedplants. In this fashion, where linkage analysis (detection of linkage disequilibrium) is possible,there are no technological barriers to extend these strategies to the vegetatively propagated cropsof interest and to genetically engineer agronomically important genes.

This CRP prepared the ground for advanced research by developing and applyingtechniques for the characterization of genetic diversity, taxonomic and phylogeneticrelationship, cultivar identification, and in exceptional cases, the preparation of preliminary coremaps in banana and yam. The techniques and markers can be used directly to continue researchtowards mapping and characterization of mutations and agronomically interesting traits invegetatively propagated crops.

Results- Molecular markers have proven potential for identifying genetic variability in vegetativelypropagated crops. Therefore this successful technology and its use should be continued.- The multiplicity of DNA marker technologies should be maintained and expanded if requiredby a specific problem, as each technology (RAPD, RFLP, AFLP, DAF, STMS, RAMPO) hasalready provided useful results in a range of crops. Despite differences in cost, reliability, andneed for prior sequence information, no single technique provides clear-cut experimentaladvantages.- The network established in the present CRP has been effective and valuable, and should bemaintained and strengthened either through organizational (future CRPs, other grantingagencies, national programmes, institutional sources) or other means (e-mail, germplasmexchange, collaborative projects, exchange of personnel).

A final technical document is now being prepared for publication.

Regional Planning Workshop of Principal Investigators on Drought Tolerance. CentreRégional de la Recherche Agronomique du Rif, Station Centrale des Radioéléments,Tangier, Morocco, 12-16 May 1997.

Plant breeders and plant physiologists nominated as principal investigators on droughttolerance from Algeria, Ethiopia, Kenya, Madagascar, Morocco, Sierra Leone, Sudan andTanzania participated. During the workshop the first year's programme was planned fordevelopment and evaluation of drought tolerant mutant germplasm of cereals and legumes. Lecturers from ICRISAT and IITA participated in the workshop. It was also agreed that eachcountry will focus activities on improvement of drought tolerance of one crop only withregional relevance: Algeria (barley), Ethiopia (chickpea), Kenya (wheat), Madagascar(groundnut), Morocco (durum wheat), Sierra Leone (maize), Sudan (groundnut) and Tanzania

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(barley). It was agreed that a multidisciplinary approach is needed and that breeders shouldtherefore collaborate closely with agronomists, physiologists and soil scientists in order tounderstand specific drought problems and establish drought selection sites for reproducibleresults. Trait-based selection for drought tolerance should be given high priority focusing on afew selected traits. Suitable selection techniques developed in International AgriculturalResearch Centres and other institutes should be adapted to national programmes to facilitate theidentification of drought tolerant mutants.

15th IAEA/FAO Interregional Training Course on Advances in Plant MutationTechniques. Agriculture and Biotechnology Laboratory, Seibersdorf, 20 May to 27 June1997.

26 participants from Algeria,Argentina, Armenia, Belarus, Bulgaria,China, Cuba, El Salvador, Ethiopia, Ghana,India, Islamic Rep. of Iran, Kenya, Rep. ofKorea, Myanmar, Nigeria, Pakistan, Peru,Poland, South Africa, Sri Lanka, Sudan,Tunisia, Uganda, Uruguay and Vietmanwere trained in basic aspects of genetics andbreeding with special regard to theirapplication to mutation induction andselection. Advances in in vitro techniquesand molecular techniques were covered inrelation to their use combined withmutation techniques. The course wasplanned and implemented to cover majorproblems and technical aspects of plantimprovement in both seed and vegetatively

propagated crops. In addition to lectures bythe staff of the Plant Breeding & GeneticsSection/Unit, lectures were also given byDr. A. Ashri and Dr. U. Lavi (Israel), Dr. S.Daskalov (Bulgaria), Dr. J. Dolezel (CzechRepublic), Dr. Z. Kaczmarek (Poland), Dr.G. Kahl (Germany), Dr. A. Lebeda and Dr.Z. Ohnoutka (Czech Republic), Dr. H.Brunner, Dr. A. Micke and Dr.Schiessendopler (Austria), Dr. I. Szarejko(Poland) and Dr. A.M. van Harten(Netherlands). Special thanks to thesepersons for the time and effort they investedto make this course a success.

Third and final RCM of the FAO/IAEA Coordinated Research Project on 'InducedMutations for Sesame Improvement'. University, Bangkok, Thailand, 6-10 April 1998.

16 participants from Australia, Bangladesh, China, Egypt, India, Israel, Kenya, Pakistan,Republic of Korea, Sri Lanka, Thailand Turkey, Uganda and USA attended this meeting whichwas held at the Kasetsart University in Bangkok, Thailand. The participants presented the finalachievement on their individual projects and formulated conclusions and recommendations onthe use of induced mutations for sesame improvement.

The main characters for improvement were determined, and for each of them a detaileddescription was given. Wherever possible the trait was split into components andrecommendations were given for the mutagen treatment and selection criteria. The maincharacters described are yield potential, harvest index, seed quality, oil quality, diseaseresistance, shatter resistance, uniform maturity and male sterility. Selection criteria weredefined for all these characters.

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The use of induced mutations has proven to be a promising technique for the geneticimprovement of sesame since many participants reported obtaining several mutants withdesirable characters like indehiscence, determinate growth, increased seed yield, diseaseresistance and water logging resistance. In some, they reached the stage of regional or nationalyield trials for their mutant lines prior to possible release of new varieties. For example theparticipant from the Republic of Korea, Dr. C.W. Kang, recently officially release a mutantderived sesame variety 'Pungsankkae' which was obtained after crossing of a local Koreanvariety with Dr. Ashri's determinate mutant 'dt-45' from Israel. Future release of new mutantvarieties is expected in the coming two years in countries such as Bangladesh, Egypt, Pakistan,Rep. of Korea. A local demonstration field was organized with accessions of mutant lines fromthe various participating countries. Observations, plant development and breeding objectiveswere discussed in the field. A wide variety of confirmed mutant lines was produced in thisproject and a database of these will be organized at the Plant Breeding and Genetics Section. Itwas recognized that exchange of useful germplasm was and remains very valuable and shouldbe encouraged as much as possible. An assessment was made of the achievements andrecommendations were made for future coordinated research efforts. A final technicaldocument is being prepared for publication.

D. STATUS OF EXISTING CO-ORDINATED RESEARCH PROJECTS

Induced Mutations in Connection with Biotechnology for Crop Improvement in LatinAmerica

The project started in 1994 followingthe recommendation of plant breeders andgeneticists involved in applications ofmutation and related biotechnologies forcrop improvement. The first Research Co-ordination Meeting was held in October,1994 in Guatemala and concentrated onchoosing the best approaches to solvebreeding problems in seed and vegetativelypropagated crops. The second RCM washeld in October, 1996 and was devoted toevaluation of results coming fromdevelopment of new mutated generationsand characterization of previouslydeveloped mutants with desired traits. The final RCM is planned for 1998. At the moment the programme has 13participants from Bolivia, Brazil, Chile,

Colombia, Cuba, Ecuador, Guatemala,Mexico, Uruguay and Venezuela. Increasedcrop productivity has become the highestpriority in most countries of the LatinAmerica Region and among factors limitingthis, the most important and most commonare: soil aluminium toxicity, salinity,drought, lack of plant available phosphorous in soil but also disease andpest susceptibility. Modern biotechnology,including induced mutations, offers anenormous possibility to breed desiredvarieties in a relatively short time. Additionally, both these techniques makeavailable the breeding of some vegetativelypropagated crops, which until now wereimproved mainly on a selection basis fromnatural or cultivated populations.

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In vitro Techniques for Selection of Radiation Induced Mutants Adapted to AdverseEnvironmental Conditions

The first RCM of this CRP was heldin 1994 in Vienna, and the second in Cairo,Egypt, in 1996. It is planned to hold the 3rd

RCM in Shanghai, China in August, 1998. At the moment the project has 10participants from Bangladesh, China,Colombia, Egypt, Ghana, India,Netherlands, Pakistan, Peru and the USA.

The participants in this CRP areworking on the improvement of potato,sweet potato, garlic, sugarcane, pineappleand alfalfa by combining in vitro techniqueswith induced mutagenesis to select for

resistance to salinity, freezing, heat,drought, and water-logging depending uponthe particular adverse condition prevailingin their country. They reported results ofradio-sensitivity tests on in vitro culturedplant material such as micropropagatedplants and organogenic or embryogeniccallus cultures. In addition, reports werepresented on the modifications of culturemedia required to regenerate and multiplylocal varieties and to carry out in vitroselection for specific stress conditions.

Induced Mutations and Other Advanced Technology for Production of Crop MutantsSuitable for Environmentally Sustainable Agriculture

At the moment the CRP has 20 participants from Belgium, Bulgaria, Brazil, Canada, China,Costa Rica, Japan, Pakistan, Poland, Sri Lanka, Turkey, UK and the USA. The second RCMwas held in San Jose - Heredia, Costa Rica in 1995, and the final RCM is planned for August,1998. At the beginning of the project it was recommended that breeders should pay moreattention to the proper plant "ideotype" for each species being bred for sustainable cropproduction. More elaboration is needed on the proper characters related to earliness, vigor,abiotic stress tolerance, and on characters related to suitability for inter-cropping production. Itwas also pointed out that the use of doubled haploids (DH) needs to be increased in all breedingprogrammes as it has become very clear that the production of DH lines is capable of speedingup variety production in areas of sustainable agriculture. It was felt that rapid screeningtechniques are the key to plant improvement when developing mutants for production understress conditions. It was concluded that there was a clear need for cheap, reliable co-dominantmolecular marker systems. PCR clones that are species- or genome-specific (i.e. micro- or mini-satellites) should be developed, and the technology for their development should be madecheap, reliable and easy to utilize in most breeding programmes. Information about past andexisting varieties of all crops and ornamentals produced by induced mutations was requested.

Radio-actively Labeled DNA-probes for Crop Improvement

The first RCM under this programme was held in Vienna in 1995, and the second RCMwas held in San José, Costa Rica in 1997. At the moment the programme has 8 participantsfrom Costa Rica, Germany, UK and the USA.

Recent advances in DNA-based technologies offer the opportunity for molecular genetictechniques to positively affect plant breeding and genome diversity characterization indeveloping countries. The use of linked DNA markers makes possible the large scaleapplication of indirect selection for important agronomic traits. This project helps to fosterinternational co-operation for transferring modern biotechnology to developing countries with

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the active participation of leading laboratories.An important activity in this project is the promotion of probes and primers including:

a) RFLP anchor probe sets for i) wheat, barley, pearl and foxtail millet, for ii) maize, rye,and rice, for iii) sorghum, and for iv) mungbean, cowpea, common bean and soybean.b) Primers for mapping and fingerprinting like arbitrary primers which can be used ingenetic mapping and marker-assisted breeding, taxonomy and systematics, differentialgene expression. The CRP participants recommended that distribution of Operon primersis limited to qualified developing countries with an appropriate level of technicalcompetency to successfully utilize such primers. A set of 10 conventional 8-mer primersis available for distribution upon request. Other more specialized primer sets (e.g. 11-merand 12-mer mini-hairpin primers) can also be made available.c) Primers for yam, chickpea, banana, plantain and phytopathogenic fungid) Primers for virus detection.

(for more information on this project, see the WWW page on the Internet under the followingURL: http://www.iaea.or.at/programmes/d2/radprobe.htm)

Improvement of New and Traditional Industrial Crops by Induced Mutations andRelated Biotechnology

The application of induced mutations and related biotechnologies to oil crop and fiberplant improvement programmes is the major emphasis of this CRP. It was established in early1995 following recommendations of several expert consultancies to include both knownindustrial crops and potential industrial crops in the process of domestication. At presentresearch activities of 16 participating institutes from Bangladesh, Brazil, Canada, China,Germany, Greece, Hungary, India, Pakistan, Spain, Turkey and US, are being co-ordinated. Thecurrent research related to improvement of oil crops as industrial crops is oriented towards thefollowing objectives:a) Development of enhanced germplasm of sunflower, linseed, soybean, rapeseed, Cuphea,

borage, cotton and jute for traditional and non-traditional, industrial and non-industrialuse. This work will be accomplished by using induced mutations, natural geneticvariants, engineered genes, classical breeding approaches and biotechnological methods;

b) Development of mutagenic approaches for additional species and screening proceduresfor agricultural and industrial requirements;

c) Development of suitable genotypes adapted to new areas and for new needs - agriculturaland industrial;

d) Assessment and demonstration of the potential of induced mutations to affect criticalsteps in various biosynthetic pathways leading to modifications in oil quality and othermetabolites;

e) Enhancement of regional and interregional co-operation in the area of inducing mutationsby radiation and other means to obtain faster breeding progress.

The Second Research Co-ordination Meeting (RCM) under the FAO/IAEA Co-ordinatedResearch Programme was held in Giessen, Germany, 30 June - 4 July, 1997. It was attended by17 participants, from Bangladesh, Brazil, Canada, China, Germany, Greece, Hungary, India,Pakistan and USA, and a representative of FAO who acted as a scientific co-secretary of themeeting. The excellent local arrangements organized by Prof. Wolfgang Friedt and his team aregreatly acknowledged.

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Recommendations made during the meeting regarded cotton, jute, soybean, sunflower,groundnut, oilseed Brassica, Cuphea, meadowfoam, flax and false flax. All agreed on the needfor germplasm exchange and to follow international safety guidelines. Mutation inductionprotocols were optimized for different species; improved germplasm was developed by inducedmutations, intra- and inter-specific hybridization and genetic transformation; molecular markersfor marker-assisted selection were developed; genes were cloned (e.g. for fatty acidbiosynthesis).

Cellular Biology and Biotechnology Including Mutation Techniques for Creation of NewUseful Banana Genotypes

This CRP was established in 1994,following the initiative of the World Bankon the Banana Improvement Project (BIP),to join efforts on the improvement of one ofthe most important crops in manydeveloping countries. The general objectiveof the project is to integrate radiationinduced mutations, in vitro culture andmolecular genetics methods into theconventional breeding of banana forgeneration of desired variation such asdisease resistance, dwarfism and earliness. In January 1996, Belgium, which has beenan important contributor to theimprovement of banana and plantainresearch in developing countries, joined thisCRP through two unique institutes: theLaboratory of Tropical Crop ImprovementK.U. Leuven which concentrates on bananagenetic manipulation and the Plant

Pathology Unit, Faculty of Agronomy,Gembloux (F.S.A.Gx.) which deals morewith banana diseases. The researchactivities proposed in this CRP are in linewith the research priorities established bythe Belgian Administration forDevelopment Corporation (BADAC).The following countries in addition toBelgium, are participating in the project:Brazil, Cameroon, Czech Republic,Colombia, Cuba, Ecuador, France,Germany, Guyana, Honduras, Israel,Malaysia, Philippines, South Africa,Tanzania, Uganda, USA, Vietnam andZaire.International Organizations:IITA (Nigeria), INIBAP (InternationalNetwork for Improvement of Banana(France).

E. NEW CO-ORDINATED RESEARCH PROJECTS

Overcoming Key Constraints to Productivity and Utilization by Genetic Improvement ofUnderutilized- and Neglected Crops in LIFDCs through irradiation and relatedtechniques.

Background:

Food availability per capita in 'LowIncome Food Deficit Countries' (LIFDCs)is on the decline, mainly as a result ofpopulation increase, but also due to

environmental factors and a lack ofimprovement of local crop species. Thesedevelopments have led not only to declinein calories per capita but also to

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nutritional imbalances and deficiencies. Furthermore, population pressure andconcentration on fewer crops to meet theever increasing food demand have led toerosion of the high genetic diversity foundin traditional agriculture leading tounsustainable agricultural practices. Inaddition, production of export orientedcommodity crops such as rubber, coffee,cacao, and African oilpalm, whileimportant to the economy, has not helpedpopulations at large to secure the minimalcalorie requirements and supply them withmicro-nutrients required to avoidmalnutrition. Increased production and asecure food supply can be achieved byimproving yield of traditional cropsespecially those grown by small landholders. The strategy for increasingproduction should maximize stability ofthe farming systems by increasing yieldper unit area without over-exploiting natural resources.

Improved crop cultivars have a keyrole in enhancing food production andsecurity. During the last FAOInternational Technical Conference onPlant Genetic Resources held in 1996 inLeipzig, Germany, a Global Plan of Action(GPA) was adopted by the Member Stateswhich included recommendations onspecific priority activities for theutilization of Plant Genetic Resources. These activities include: (1) expanding thecharacterization, evaluation and number ofcore collections to facilitate use; (2)increasing genetic enhancement and base-broadening efforts; (3) promotingsustainable agriculture throughdiversification of crop production andbroader diversity in crops; (4) promotingdevelopment and commercialization ofunderutilized crops and species; (5)supporting seed production anddistribution; and (6) developing newmarkets for local cultivars and “diversity-rich” products. The GPA calls for

international action to contribute to thesepriority activities.

Only a few crops produce themajority of the world’s food, yet manyneglected and underutilized crops areextremely important for food production inlow income, food-deficit countries. Neglected crops are those which aretraditionally grown in their centres oforigin or centres of diversity by farmers,where they are still important for thesubsistence of local communities. Thesecrops continue to be maintained by socio-cultural preferences and traditional uses. Examples include the Andean root andtuber crops such as ahipa, aracacha, maca,yacon, ulluco, the minor millets such asPanicum, Paspalum and Digitaria speciesand many leafy vegetables. A few of thesecrop species may be distributed morewidely, but tend to occupy special nichesin the local ecology and in production andconsumption systems. Neglected cropsremain inadequately characterized, anduntil very recently have been largelyignored by research and conservation. Underutilized crops, are those whichwere once more widely grown such asbambara groundnut, buckwheat, sesameand safflower, but are now falling intodisuse for various agronomic, genetic,economic and cultural factors. Farmers arelosing these crops because they are lesscompetitive with improved major cropspecies.

Many of these neglected andunderutilized crops are locally welladapted and constitute an important part ofthe local diet, culture, and economy;require relatively low inputs; andcontribute to high agriculturalsustainability. Traditional agriculturalresearch in developed countries hashitherto paid little attention or ignoredthese crops and consequently they haveattracted little research funding despite thefact that they are adapted to a wide range

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of growing conditions, contribute to foodsecurity, especially under stress conditions,and are important for a nutritionally well-balanced diet. Reduced public funding foragricultural research combined with thetrend towards privatization of the seedindustry further exacerbates this problem. These traditional crops often are lowyielding and cannot compete economicallywith improved cultivars of major crops,even though many of these crop specieshave the potential to become economicallyviable. A major factor hampering thedevelopment of these traditional crops isthe lack of genetic improvement and attime narrow genetic diversity for importantagronomic traits. Further constraints arethe lack of knowledge on the taxonomy,reproductive biology, and on the geneticsof agronomic and quality traits.

In many LIFDCs, traditional cropsare frequently heterogeneous land races,and often represent well adapted,domesticated germplasm. In many casesthe constraints to increased productivityand utilization of these traditional cropsare caused by specific bottlenecks such asseed shattering, indeterminate growth,uneven maturity, unsatisfactorypostharvest and end-product quality,photoperiod sensitivity, disease and insectsusceptibility and lack of abiotic stresstolerance. Genetic improvement can beachieved by various techniques such asgenetic recombination and selection,mutation induction, and appropriatebiotechnological approaches. Theapplication of mutation technology has adistinct advantage in that small geneticchanges which affect critical agronomictraits, can be induced or transferred to well

adapted and culturally accepted localcultivars. These techniques which arebeing used to complement conventionalplant breeding approaches in developedcountries are either unavailable or are intheir infancy in LIFDCs. Inducedmutations successes include: increasedstarch granule size leading to reducedcooking time in cassava (Asare et al.,1997); determinate growth and reducedshattering in sesame (Ashri, 1988;Cagirgan 1996; Maneekao 1997; Wongyaiet al. 1997); early maturity in buckwheat(MBNL 1993); black spot diseaseresistance in Japanese pear (Sanada et al.1993); drought tolerance in amaranth(MBNL 1994); semi-dwarfness in rice(Rutger, 1983); and improved shelf-life inapple (MBNL 1988). Many key traits indomestication or crop improvement arecontrolled by single or a few genes (seeAnnex 2) and can be obtained bymutations.

This CRP will focus on the geneticimprovement of a number of neglected andunderutilized species aiming to overcomekey constraints to productivity andutilization and thus stem the erosion ofcrop diversity. This project will bedeveloped through the establishment of aresearch network of partners in thedeveloping and developed Member States. The project aim will be to apply and adaptexisting technology, already developed formajor crops, to neglected and underutilizedcrops in order to improve food security,enhance nutritional balance, and promotesustainable agriculture.

Criteria to be used for the selection of crops:For a neglected and underutilized crop to be included in this project, it will be necessary

to fulfill most of the following criteria:• Significant contribution to nutrition and food security• Availability of local and other germplasm

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• Evidence of local breeding activities• Key constraints resolvable through mutation techniques• Existence of regional/international networks• Economic viability• Acceptance by growers and consumers

Potential crops to be included in this project:Cereals and pseudo-cereals• Amaranth (Amaranthus spp.)• Quinoa (Chenopodium quinoa)• Teff (Eragrostis tef)• Buckwheat (Fagopyrum esculentum)• Minor millets (e.g. Panicum spp., Paspalum spp., Digitaria spp.)

Grain legumes and oil seeds• Bambara groundnut (Vigna subterranea)• Jack bean (Canavalia ensiformis)• Niger or noug (Guizotia abyssinica)• Sesame (Sesamum indicum)• Grass pea (Lathyrus sativus) Root and tuber crops• Andean root and tuber crops (e.g. Oxalis tuberosa, Ullucus tuberosus, Lepidium meyenii,

Polymnia sonchifolia)• Yam (Dioscorea esculenta)• Taro (Colocassia esculenta), Yautia (Xanthosoma sagittifolium)

Fruits and vegetables• Tropical fruits [e.g. Durian (Durio spp.), Rambutan (Nephelium lappaceum)]• Peach palm (Bactris gasipaes)• Pitaya (Hyloscereus ocamponis, Cereus peruvianus)• Okra (Abelmoschus esculentus)• Minor solanums (e.g. Solanum quitoense, S. gilo, S. macrocarpon)• Bitter gourd (Momordica charantia)

Countries:For Research Contracts a priority will be given to participants from LIFDCs.

However, participants from other developing countries will not be excluded if crops ortechnology are concerned with potential benefits for the LIFDCs.

Overall objective:To improve food security, enhance nutritional balance, and promote sustainable

agriculture in LIFDCs.

Specific objective:To overcome major constraints to productivity of neglected and underutilized crops by

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genetic improvement, in order to enhance economic viability and sustain crop speciesdiversity, and in future to benefit small farmers.

Expected outputs:1. Identified and prioritized major constraints to the development of a number of locally

adapted neglected and underutilized crops.2. Establishment of a network for research cooperation, and germplasm conservation and

use.3. Constraints to development overcome by genetic improvement techniques, including

selection and cross-breeding, mutation techniques, and appropriate biotechnologicalapproaches.

4. Availability of enhanced evaluated germplasm for a number of investigated neglectedand underutilized crops.

5. Developed protocols for genetic improvement techniques of individual under-utilizedcrops.

6. Technology transferred and dissemination of information through publications.

General Assumptions:· Interested and competent partners.· Key constraints to productivity and utilization of the neglected and underutilized crops are dueto traits controlled by one or a few genes.· The genes required are either available in existing germplasm locally or in collections or canbe induced through mutation technology.· The necessary breeding and mutation technology is currently available for adoption.· The required breeding efforts will be carried out locally.· Adequate local support and Agency inputs provided.

To apply for a contract/agreement in this project, please complete a Proposal for ResearchAgreement or Research Contract Proposal and return it to the IAEA’s research ContractsSection.

Mutational Analysis of Root Characters Related to Drought Tolerance to Sustain CropProduction in Arid and Semi-Arid Zones

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Background:Most of the world's hungry live in

countries that are classified as low-incomeand food-deficit (LIFDCs). More than 80countries are considered in this category andover half of them are in Africa. Most of theLIFDCs are located in areas where droughtis a major factor limiting crop production.Critical evaluation of progress in plantbreeding over past decades hasdemonstrated genetic improvements in yieldunder both favourable and stress conditions. The yield improvement under droughtstress resulted partly from geneticimprovement of yield potential and partlyfrom improvement of stress resistance orgrowth duration. Genetic and plantphysiological investigations have indicatedthat several mechanisms lead to an increaseof drought tolerance in cereals. Amongthem are: maintenance of high leaf waterpotential under stress, deeper root growth,higher root density, higher root mass,stomatal control over transpiration, osmoticadjustment, proline accumulation, highepicuticular wax load and cellularmembrane stability. All these mechanismsare usually polygenically inherited and assuch are difficult to apply as selectioncriteria. It is also considered that the use ofyield as a selection index under stress isinefficient and usually limited to aparticular drought condition. It is thereforerecommended to develop trait-basedselection procedures for improvement ofdrought tolerance.

There is no doubt that droughttolerance depends on desired characters ofthe stem and root system. Although manyinvestigations were carried out on variousstem characters in relation to abiotic stresstolerance, investigations on root systemcharacters are relatively limited. The simplefact that roots grow in soil and as such areinvisible is the greatest inhibitor toadvancement of knowledge on root systemsand particularly about their genetics. For

the same reason there is very limitedknowledge about the level of geneticdiversity in root systems (only about 30plant genera have been investigated for theevaluation of genetic variation of roots). Recent emphasis on the genetic diversity ofroot system has concentrated on the needfor increased drought, salt, acid soiltolerance or nutrient uptake. Unfortunately,the great environmental component of rootsystem variability is an additional factorcomplicating a clear genetic conclusionfrom such investigations. In contrast to theplant shoot where many morphologicalcharacters such as shape and colour offlower, leaf or culm were well describedand genetically investigated long ago, inroots, there are only a few known characterswhich could be used as genetic markersfacilitating genetic analysis ofagronomically important physiologicaltraits. In addition, where variation was discovered, it is often polygenic in nature.

Against this background geneticistsand plant breeders are now using mutationtechniques to generate monogenicallyinherited root morphological markers. Thisis now more realistic as somenondestructive methods of rootinvestigation, including hydro- andaeroponics and computer image analysis,have been developed. Also very promisingresults have been obtained on the modelplant species - Arabidopsis. These studieshave shown that numerous important rootcharacters can mutate after treatment ofseeds with physical and chemical mutagensor as a result of insertional mutagenesis, andthat large numbers of already developedroot mutants are monogenically inherited.Similarly, preliminary results obtained inbarley and maize indicate a high frequencyof root system mutants in mutatedgenerations. It is therefore now possible toinduce and develop a germplasm collectionof root system mutants and to initiatemutational analysis of both morphological

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and physiological characters; therebyestablishing suitable breeding programmesfor increased tolerance to drought or otherstresses, with an improved root system asthe main component. The latestdevelopments in biotechnology willfacilitate implementation of such aprogramme through rapid homozygotizationof mutants and molecular marker assistedselection of desired genotypes followed bycharacterization, isolation and cloning ofthe most important mutated genes.

Objective:To make mutational analysis of some

of the essential characters related to rootformation and functioning, theircontribution to crops' tolerance to drought,and the effects of the relevant mutated rootsystem characters including their variousphysiological manifestations and linkages.

Expected outputs:This project will lead to development ofmutant lines with desired root types and soilpenetration dynamics. Furthermore, thesemutant lines will be geneticallycharacterized and a strategy will bedeveloped for the use of root systemmutants in cross breeding programmes fordrought tolerance.Action plan:

- Induce mutants with altered rootsystem characters- Select and evaluate promisingmutants with desired root types andaltered soil penetration habits.- Conduct genetic analysis of theselected mutant lines.- Incorporate root system mutants inexisting crop improvement and crossbreeding programmes in the specificplants.

If you are interested in applying for acontract/agreement in this programme,

please complete a Proposal for ResearchAgreement or Research Contract Proposaland return to the IAEA's Research ContractsSection. The final selection of the crops,objectives and approaches will be madeafter the proposals are evaluated.

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F. ACTIVITIES AT THE PLANT BREEDING UNIT, SEIBERSDORF

The Plant Breeding Unit of theFAO/IAEA Agriculture and BiotechnologyLaboratories at Seibersdorf providestechnical support to the Joint FAO/IAEADivision's programme in the area of plantbreeding and genetics. This support involvesthree major components;- research and development in advancedbreeding techniques using nuclear andbiotechnological methods in crops such as

banana, plantain and rice;- training of individual fellows and groupsof scientists from developing MemberStates;- providing mutation induction services andtechnical advice to scientists in MemberStates.

The current activities cover the following:

Induction and verification of autotetraploids in diploid (Musa acuminata) by in vitrotechniques.

Flow cytometry and stomatacharacteristics were used for screeningploidy levels in a large population of in vitroinduced autopolyploids of the Musaacuminata breeding clone SH-3362.Culturing shoot tips in liquid mediumsupplemented with 5.0 mM colchicine for 48hours or 30 µM oryzalin (3,5-dinitro-N4,N-dipropylsulphate) for seven days, both incombination with 2% (v/v) DMSO, resultedin a high (23 % and 29 %) frequency of non-chimeric tetraploids in the fourth vegetative

generation. Although mixoploidy persistedin subsequent cycles of vegetativepropagation, tetraploids as identified by flowcytometry remained solid non-chimericduring two more cycles. Theseautotetraploids were propagated for fieldtesting. A rough pre-selection of regeneratedV4 plants based on their stomatacharacteristics resulted in a population inwhich only 56 % of the plants were solidtetraploids.

Histology of somatic embryo initiation and organogenesis from rhizome explants of Musaspp.

Bananas and plantains are important staplefood crops for people living in tropical andsub- tropical countries. Declining yields dueto the spread of virulent diseases such asBlack Sigatoka, Fusarium wilt and BananaBunchy Top Virus, has resulted in increasedefforts to genetically improve this crop.However, conventional breeding of Musacultivars remains a difficult endeavorbecause of high sterility and polyploidy.

Plant tissue culture techniques canpotentially overcome some of the factorslimiting traditional approaches to banana andplantain improvement. These techniquesenable plants to be regenerated from normal

and genetically modified cells and tissues inan efficient way under sterile conditions. Thegeneration of genetic variation by inducedmutations or genetic transformation is asingle cell event. Therefore treatment ofmulti-cellular cell initials leads to theformation of chimeras. Genetically alteredand non-altered cell-lineages compete duringthe proliferation and growth of multi-cellularinitials and lead to the formation of

mericlinal or sectorial chimeras. Theseare not manifested phenotypically andrequire repeated subculture to rescuegenetically modified homohistont tissueprior to mass-screening and/or selection fordesirable traits. The genetic improvement of

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Musa by in vitro mutation breeding orgenetic engineering requires the efficientregeneration of genetically modified singlecells into complete plants.

The origin of somatic embryos derivedfrom rhizome explants of triploid Musa c.v.Grand Naine, was the subject of histologicalstudies during different phases ofontogenetic development. Our histologicalobservations revealed that the embryogenicmass and somatic embryos developed inmost instances from several morphologicallycompetent adjacent cells while theoccurrence of single cell origin in vasculartissues of rhizome explants of Musa was less frequent. The majority of somaticembryos showed normal root formation andconsisted of highly vacuolated cells in thepoorly structured shoot apex.

Preliminary studies on the use ofmutation induction and anther culture inthe improvement of Eragrostis tef

Tef is the single dominant crop grownwidely amongst the indigenous andintroduced field crops of Ethiopia. It isconsidered to be well adapted to drought andwaterlogged conditions. Lack of adequategenetic improvement of the present cultivarsand agronomic problems such as lodging and leaf rust are the major causes for lowyield. Of the more than 2000 germplasmmaterials collected so far, none is resistant tolodging. Therefore, induced mutationbreeding has been considered as one of thealternatives.

Assessment of optimal gamma ray and EMSdoses

Radiosensitivity of seeds to gammarays was genotype specific. Suitableconcentrations for treating tef seeds withethylmethane-sulphonate (EMS) werebetween 0.6% and 0.9% with the appliedchemomutagen treatment regime.

Anther cultureSignificant genotypic differences were

noted in the callus formation in antherculture. A very high proportion (88-97 %)of the spikelets cultured produced calli. Nosignificant difference was obtained amongthe four genotypes used and the responsewas found to be high both from gamma raytreated and untreated spikelets. The youngspikelets of tef could be used as an excellentsource of explant to induce callus for varioustissue culture purposes. Studies still need tobe done to determine if spikelets are apossible source of explant for the productionof doubled haploid plants.

Regeneration of Ensete ventricosum through somatic embryogenesis andadventitious budsIn southern and south-western Ethiopia,Ensete ventricosum is grown as an importantstarchy, staple food crop, supporting the dietof a quarter of the Ethiopian population. Dueto difficulty in germinating seed and the longvegetative period, breeding ensete isextremely difficult. Adventitious buds andsomatic embryos were induced from callusderived from corm tissue and cultured onMurashige and Skoog’s (MS) basal mediumsupplemented with benzylaminopurine(BAP) or 2iP. Elongation of somaticembryos was achieved on the same mediumand rooting was induced on half strength MSbasal medium supplemented with IBA. Nophenotypic variation was observed amongmore than 200 potted regenerants.

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Assessment of genetic diversity in three African populations of malabar spinach (Basellaspp.)

Malabar spinach (Basella spp.) is an important leaf vegetable in the human diet of WestAfrica. The nuclear genome size, the ploidy state, the degree of genetic variation present in thethree African populations of Malabar spinach (Congo domesticated, Sri Lanka, vegetativelypropagated) were investigated using chromosome counting, flow cytometric techniques andRAPDs (Random Amplified Polymorphic DNA) analysis.

The sexually propagated samples, i.e. the two populations named Sri Lanka and Congodomesticated, showed the same ploidy level (2n=36). These two populations also gave similarresults with a total DNA content of 6.83 pg (2C). The third population, vegetatively propagated,showed a lower DNA content of 5.73 pg. In the vegetatively propagated plants complexmixoploidy was observed with chromosome numbers ranging from 36 to 48. Fragment profilesobtained with the RAPD primers were scored manually as present (1) and absent (0). This methodof scoring elucidated an average of 11.9 loci per primer. From a total of 296 amplificationsproducts scored, 31% were found polymorphic. Of the 94 primers, 35 detected no polymorphismalthough they successfully amplified a range of monomorphic bands. The average number ofRAPDs detected per polymorphic primer was 3.7.

The low level of polymorphism detected among the three African populations of Malabarspinach, may reflect the tendency of inbreeding crop species to develop towards homozygositywhen compared to an obligate outbreeder. It can also be symptomatic of a narrow genetic base ora loss of diversity when introduced from Asia during the early colonial times in Congo, whereplants probably derived from the few seeds or cuttings.

Supporting services for application of mutation techniques in Member States

A radiation treatment service is provided to FAO and IAEA Member States to supportnational crop improvement programmes. Seed and vegetative plant structures were received andtreated with 60Co gamma rays and fast neutrons.

RADIATION SERVICE STATISTICS (1996-1997)No. of treated samples 675No. of treated species 36No. of treated cultivars 159No. of recipient Member States 30Seed samples 674Vegetatively propagated samples 1No. of 60Co gamma ray treatments 515No. of fast neutron treatments 160

G. PUBLICATIONS

1. Afza, R., Van Duren, M. and Morpurgo, R. 1996. Regeneration of Ensete ventricosum throughsomatic embryogenesis and adventitious buds. Plant Cell Reports 15: 445-448.

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2. Chen, Q. F., Afza, R. and Brunner, H. 1996. Induction of Somatic Embryos and Regeneration ofPlantlets from Rice Roots. Acta Agriculture Nucleatae Sinica 10(4), 216-220.

3. Gwanama, C. and Nichterlein, K. 1996. Leaf yield components and nutritional quality ofZambian pumpkin (Cucurbita L.) land races. Cucurbits towards 2000. Proceedings of the VIthEucarpia Meeting on Cucurbit Genetics and Breeding. Malaga, Spain, 28-30 May, 1996.Gomez-Guillamon, M.L., Soria, C., Cuartero, J., Tores, J.A., and Fernandez-Munoz, R. Malaga:Estación Experimental "La Mayora", C.S.I.C.: 134-140.

4. Nichterlein, K., Van Zanten, L., Maluszynski, M., Ahloowalia, B., and Weck, E. 1996. FAO/IAEA Programmes on improvement of oil crops by induced mutations and relatedbiotechnologies. Eucarpia Section Oil and Protein Crops. Symposium on Breeding of Oil andProtein Crops, 5-8 August 1996, Zaporozhye, Ukraine Institute of Oilseed Crops of theUkrainian Academy of Agricultural Sciences: 159-164.

5. Van Duren, M., Morpurgo, R., Dolezel, J. and Afza, R. 1996. Induction and verification ofautotetraploids in diploid banana (Musa acuminata) by in vitro techniques. Euphytica 88: 25-34.

6. Weck, E., Grasso, G. 1996. How similar are plant telomeres? Maize Genetics CooperativeNewsletter 70: 66.

7. Use of mutation techniques for improvement of cereals in Latin America. IAEA-TECDOC-859.Vienna (1996). This document contains scientific results obtained under a regional CRPincluding promising mutants of important crops for the region. Copies can be obtained bywriting to the Section.

8. Ahloowalia, B.S. Maluszynski, M., Nichterlein, K., van Zanten, L. and Weck, E. (1997) Induced mutations and in vitro culture techniques for the improvement of horticultural plants. Chapt. 7, Proc. 2nd Intnl. Crop Sci. Congress. New Delhi (in press)

9. Polok, K., Szarejko, I, Maluszynski, M. 1997. Barley mutant heterosis and fixation of F1-performance in doubled haploid lines. Plant Breeding 116: 133-140.

10. De Moura, D., Zapata-Arias, F.J., Ando, A. and Tulman Neto, A. 1997. Plant regeneration fromprotoplasts isolated from primary calli using mature embryos of two Brazilian rice cultivars.Euphytica 94:1-5.

11. Improvement of basic food crops in Africa through plant breeding including the use of inducedmutations. IAEA-TECDOC-951, Vienna (1997). A final document from a CRP funded by theItalian Government. It contains reports, conclusions and recommendations from the participantsregarding the development of improved varieties in staple food crops in Africa. Copies can beobtained by writing to the Section.

12. Bayoud Disease of Date Palm. Document issued by the IAEA Technical CooperationDepartment, Vienna (1997). Report of consultants meeting organized by the Joint FAO/IAEADivision of Nuclear Techniques in Food and Agriculture, Vienna 25-29 May 1996.