A four-nucleotide base-pair deletion in the coding region of the Bowman-Birk protease inhibitor gene prevents its accumulation in the seeds of Glycine microphylla PI440956

Abstract

The Bowman-Birk protease inhibitor (BBI), an abundant soybean [ Glycine max (L.) Merr.] seed protein, is a major antinutritional factor. Nulls for the major soybean BBI have been reported in several of the wild perennial Glycine species including G. microphylla (Benth.) Tind PI440956. This perennial Glycine species does not accumulate the major BBI and the molecular basis for the absence of the major BBI in this plant introduction (PI) line is not known. We have cloned the BBI gene from G. microphylla PI440956, G. microphylla PI505188, and G. max cv. Jefferson and determined its nucleotide sequences. Analysis of the G. microphyllla PI505188 and G. max cv. Jefferson nucleotide sequences revealed a complete open-reading frame encoding the BBI. In contrast, the BBI coding region of G. microphylla PI440956 contained a frameshift mutation that resulted in the introduction of a stop codon at the amino terminal region of the protein. Reverse transcription-polymerase chain reaction analysis revealed that the BBI gene was expressed in developing seeds of G. microphylla PI505188 and G. max cv. Jefferson, but not in developing seeds of G. microphylla PI440956. In contrast, a BBI-related isoinhibitor gene was expressed at similar levels in all three Glycine species. Our results suggest that the frameshift mutation in the BBI coding region is responsible for the absence of BBI in the seeds of G. microphylla PI440956.