Diversity of the waxy gene in cultivated and wild rice in Thailand

Rice Waxy gene encodes a granule-bound starch synthase (GBSS) for synthesis of amylase in endosperm tissue. In Thailand, there has been a lot of rice varieties with variation in apparent amylase content. Amylose content is a key determinant of the cooking and processing quality of rice (Oryza sativa). Low amylase levels are usually associated with tender, cohesive, glossy cooked rice, while higher amylase levels tend to cook dry, be fluffy and separated. The objectives of this work are as follows: 1) To determine and accumulate of microsatellite classes located in Waxy gene, and the mucleotide sequence at the junction between exon1-intron1 of the wx locus. These allelic differences are further analyzed to determine the correlation with amylase content in their endosperm of rice strain used in this study. 2) To examine gene expression at the wx locus both Waxy gene product (Waxy protein) and mRNA levels by using SDS-PAGE and RT-PCR techniques, respectively. The results obtained are as follows: 1) Among 63 strains examined, ten classes of microsatellites (or CT sub(n)) repeats) were detected (n=4, 8, 9, 10, 11, 12, 14, 17, 18 and 19). One predominant class of (CT sub(17)) repeats was observed in the samples. 2) An unique microsatellite allele, (CT sub(18)), was detected in glutinous cultivars and the cultivars with low amylase content. In additional, the microsatellite allele, (CT sub(11)), was also detected only in all the cultivars with intermediate and high amylase content. 3) Rice strains with low amylase content had the sequence GTTATA (allele Wx sup(b)) or GATATA (allele Wx sup(c)) at the putative leader intron 5’ splice site, while all rice strains with intermediate and high amylase had GGTATA (allele Wx sup(a)) Furthermore, rice strains with intermediate and high amylase could be detected by using a restriction enzyme, Accl to cleave the PCR products. 4) Based on the molecular evidence, the allele Wx sup(c) in the wx locus was responsible for amylose synthesis as well as the other alleles.