Validation of DNA sequencing of waxy gene for amylose of selected traditional rice varieties through iodine staining method
2018
Romero, M.V. | Mamucod, H.F. | Huliganga, R.C. | Corpuz, G.A. | Enriquez, J.O.S. | Millas, R.A. | Tabanao, R.A.
Amylose content (AC) is the most important factor affecting not only cooking and eating quality but also processing behavior of rice. Amylose is synthesized by the granule-bound starch synthase (GBSS) enzyme encoded by the WAXY locus in chromosome 6. The wild type allele, Wxa, is commonly found in cultivars from Indica subspecies while the Wxb allele is commonly distributed in Japonica subspecies. Base substitutions from G T in Wxb allele decrease splicing efficiency which affect the enzyme activity of GBSS and results in an endosperm with lower levels of amylose. This study determined the DNA sequence for WAXY locus for amylose content of 87 traditional rice varieties (TRVs) and validated through the iodine staining method. DNA was extracted from two-week old rice seedlings of TRVs. Polymerase chain reaction was performed in order to detect G/T base substitution and (CT)n repeats in the WAXY locus. Sequence analyses revealed polymorphism in the WAXY locus for AC showing G/T substitution SNP, and variation in (CT)n repeats. The functional nucleotide polymorphisms (FNP), G at the G/T base substitution and (CT) repeats nt 16, for intermediate to low AC were observed in 80 TRVs. A total of 75 TRVs showed similar findings when subjected to amylose analysis using iodine staining method. The difference in the genotypic and phenotypic data for some TRVs can be attributed to environmental factors leading to variation of about 6% in the AC of a certain variety. This study established an excellent match between the results of the iodine staining method for AC determination and DNA sequencing of waxy gene.
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