Establishment of a soybean (Glycine max Merr. L ... - Plant Sciences

Planta (2009) 229:279–289 287Fig. 8 Putative somatic Ds transposition in the soybean genome. F 2individuals derived from a cross between Ac-transposase parent 538-5used as male and pPTN335 event 457-13 used as female. Lanes designatedwith ‘U’ and ‘L’ refer to upper and lower leaf tissue samples.Lanes under F 2-8 are double null. Ds lanes are samples from femaleparent, while Ac lanes are male parent + and ++ lanes indicate 50 pg ofpPTN335 and pPTN398, respectively. Blot was hybridized with GUSORFDNA with transposon-based systems as the delivery vehiclefor activation tag elements allows for the launch sitesto be strategically situated within transcriptionally activeregions of the genome. Moreover, multicopy integrationsoften occur with T-DNA and some chromosomal perturbationmay be induced proximal to the site of integration(Kim et al. 2003; Ohba et al. 1995; Olhoft et al. 2004),which may impact the activation of a nearby gene.Coupling T-DNA with transposon-based systems circumventsthis potential problem given the movement of thetransposon from the original site to the new location willresult in a single copy intact integration with fewer proximalrearrangements in the genome (Marsch-Martinezet al. 2002).Limited information is available on the activity of themaize Activator in the soybean genome (Aljanabi et al.1999; Zhou and Atherly 1990). We demonstrated here putativesomatic and germinal transposition of Ds is induced inthe soybean genome upon stacking a constitutive Ac-cDNAexpression cassette, with a non-autonomous Ds allele.However, one of the drawbacks of inducing Ds transpositionwith a constitutive Ac cassette is that somatic transpositionstend to be more frequent than germinal. In addition,transposition can occur throughout plant development,which may complicate cloning of the desired tagged gene.To circumvent these potential pitfalls associated with theuse of a constitutive Ac transposase, we assembled an Accassette under the control of the Arabidopsis meiosis-speciWcDMC1 promoter (Klimyuk and Jones 1997). Transgenicsoybean events carrying this latter Ac cassette willpermit comparative studies designed to monitor germinalDs transposition frequencies induced upon stacking of theconstitutive verses meiosis-speciWc transposase expression.Clearly data gathered from such a study, monitoring theaVect of Ac expression over development and level on thefrequency of Ds transposition in the soybean genome, willbe a key factor that determines the ultimate power of thismaize two component transposon system as a tool for functionalgenomics in the crop.The limited screening of the events currently housed inthe transposon-based soybean collection has revealed thatlaunch sites established by T-DNA integration tend toreside in gene rich regions (Table 1). Analysis of genesequences across taxa provided insight into what transgeniclocus may be linked with the male sterile phenotypeobserved in event Ds82, highlighting the power of comparativegenomics brought about by the previous investmentsmade in plant genomics.Arguably one of the most critical components that willenable a transposon-tagged repository to be of value to thesoybean community is the establishment of a reliable identity-preservedstorage and distribution system. Importantly,the collection must be linked to a user-friendly searchabledatabase. To this end we developed GmGenesDB (http://digbio.missouri.edu/gmgenedb/index.php). This databasehas a set of tools to Wnd insertions in gene(s) of interest, andto retrieve all annotations related to a tagged gene. Additionalinformation of the respective soybean events housedin the repository includes foreign locus number, genotypeof the event, and phenotype observed. The ultimate goal ofthis program is to provide a resource to the soybean communityto facilitate the translation of the basic tools toapplied technologies that can be exploited for downstreamimprovements in soybean genetics.Acknowledgments This work was supported by grants providedthrough USDA/NRI Award No. 2002-35301-12195, and the UnitedSoybean Board. Additional funds provided through the National Centerfor Soybean Biotechnology and Nebraska Research Initiative.ReferencesAljanabi SM, Shibli R, Ajlouni M (1999) Insertion of maize transposableelement Ac into soybean (glycine max L. Merr.) by Agrobacteriummediated transformation method. Dirasat. Agric Sci26:226–239Allen RD, Bernier F, Lessard PA, Beachy RN (1989) Nuclear factorsinteract with a soybean β-conglycinin enhancer. Plant Cell 1:623–631Altschul SF, Madden TL, SchäVer AA, Zhang J, Zhang Z, Miller W,Lipman DJ (1997) Gapped BLAST and PSI-BLAST: a new generationof protein database search programs. 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