Genome-wide scan of the soybean genome using degenerate oligonucleotide primed PCR: an example for studying large complex genome structure

Abstract

The availability of next generation sequencing instruments has made large-scale and genome-wide sequence analysis more feasible in a wild variety of species with large complex genomes, especially crops. This report suggested an approach for characterizing large complex genomes of less-studied/orphan crops. Degenerate oligonucleotide primed PCR (DOP-PCR) is a useful tool for the survey of genomes in less-studied/orphan crops, as prior sequence information is not necessary. Here, four different degenerate primers were redesigned from previously described DOP-PCR primers. The degeneracy of these primers was increased with the addition of two more ‘Ns’. The amplified DOP-PCR products from Sinpaldalkong 2, a soybean genotype, were applied to GS-FLX and the reads from Sinpaldalkong 2 were mapped against Williams 82 as a reference (http://www.phytozome.net/soybean.php), using the Burrows-Wheeler Aligner (http://bio-bwa.sourceforge.net/). These results suggest the identification of 4 single nucleotide polymorphisms between Sinpaldalkong 2 and Williams 82 and recent duplication of the soybean genome. The sequenced reads were subsequently assembled into contigs by Newbler under default conditions. A total of 29 Sinpaldalkong 2 contigs exhibited 95% similarity and < E-100 when mega-blasted with Williams 82 reference sequences. These contigs were mapped to the soybean chromosomes and positioned as clusters within each chromosome. Most of the contigs also showed similarity with the Arabidopsis RNase H domain-containing protein, suggesting a potential way to study retrotransposons in less-studied/orphan crops. Using these modified DOP-PCR primers and GS-FLX, it is possible to obtain insight into the large complex genomes of less-studied/orphan crops.