Transposon-Based Functional Characterization of Soybean Genes

Abstract

Type II transposable elements that use a cut-and-paste mechanism for jumping from one genomic region to another are ideal for use in tagging and cloning genes. Precise excision from an insertion site in a mutant gene leads to regaining the wild-type function. Thus, the function of a gene can be established based on the mutant phenotype and the regaining of the wild-type phenotype following precise excision of the element. Heterologous type II transposable elements including the Ac/Ds system from maize, the miniature inverted repeat system, mPing from rice, and the Tnt1 retrotransposon from tobacco have been successfully applied in functional analyses of soybean genes. Although several endogenous transposable elements have been identified in soybean, evidence of an active type II transposable element in soybean was largely lacking. We have previously reported the isolation of the type II soybean transposon Tgm9 from intron II of the dihyroflavonol-4-reductase 2 (DFR2) gene of the W4 locus. Tgm9 is an active element and produces variegated flowers through somatic excision. Excision of the Tgm9 element from the progenitor cells of flower buds results in genotypes with purple flowers that are known as germinal revertants. The element was discovered from a commercial soybean cultivar, and the line carrying the element was termed T322. The T322 genome contains only one active Tgm9 copy in the W4 locus. In a recent study, the utility of Tgm9 was assessed by studying a set of random germinal revertants. The new mutations created following excision of Tgm9 from DFR2 were evaluated using a transposon display assay. This study revealed that Tgm9 transposes to all 20 soybean chromosomes from its original site in the DFR2 gene. Although Tgm9 exhibited preferential transposition to a few genomic regions, across the entire genome 25.7% of the new Tgm9 mutants were detected in exon or intron sequences. Thus, Tgm9 is a suitable endogenous type II transposon to generate an indexed insertional mutant collection for functional characterization of most of the soybean genes.