Abundant Variation of Gene in Yunnan Rice Landraces and Molecular Characterization of a Novel Allele

摘要

Waxy is the key gene that controls the apparent amylose content (AC) in rice (Oryza sativa L.) endosperm. In this study three allele-specific polymerase chain reaction (AS-PCR) primer sets for Wxmq and Wxop/hp were developed that provided new markers for determining and selecting targeted Waxy alleles. The AC determination and AS-PCR results indicated abundant AC variation and extremely rich Waxy allelic diversity in Yunnan rice landraces such that 70 Waxy alleles were detected in 404 landraces. For nonglutinous rice, association analysis showed that only two of the seven Waxy functional single nucleotide polymorphisms (SNPs) were significantly correlated to AC variance. The intron 1 splice site (In1) explained 3.45% and all seven sites explained 9.79% of the total AC variance, respectively, which were much less compared to previous studies. A novel Wxzm allele with a nonsynonymous C to T transition at E10ZM site located at 2134 bp of exon 10 was characterized from a glutinous landrace Zimi. This transition led to a Leu to Phe substitution at the glycosyl transferases domain of granule-bound starch synthase I (GBSSI), which was highly conserved in Poaceae crops. Protein analysis showed that Zimi had similar endosperm GBSSI accumulation as other nonglutinous landraces but its activity was dramatically decreased, indicating that this Leu residue was critical for the glycosyl transfer activity of GBSSI. The characterization of Wxzm revealed a new mechanism for rice glutinous formation and suggested multiple origins of the rice glutinous phenotype.