A PCR-based method to test for the presence or absence of β-conglycinin α′- and α-subunits in soybean

The soybean cultivar Yumeminori, which lacks the alpha'- and alpha-subunits of beta-conglycinin, carries both naturally occurring and induced mutations. While the cause of the natural mutation resulting in the alpha'-subunit deficiency has been determined, the induced mutation in the CG-2 gene encoding the alpha-subunit has not been characterized at the molecular level. In this study, we identified a four base pair insertion in the first exon of CG-2, which introduced a premature stop codon. The insertion co-segregated with the lack of alpha-subunit, indicating that this mutation is the cause of the alpha-subunit deficiency. A multiplex PCR method of testing for the presence or absence of alpha'- and alpha-subunits was developed based on the sequences of mutated and wild-type alleles. This PCR-based test was also capable of detecting the presence of wild-type genes when Yumeminori DNA samples were contaminated with wild-type DNA at levels of 0.2% or greater. Thus, this method will be useful both for marker-assisted selection in soybean breeding programs, and for seed purity tests in food industries.