Identification of mutations in OASA1 gene from a gamma-irradiated rice mutant population

Abstract

One thousand three hundred and fifty homologous rice M8 mutant lines resistant to 5-methyltryptophan, a tryptophan analogue, were selected from embryogenic calli irradiated with 30120 Gy of gamma rays. The mutant rice would be compromised in tryptophan homoeostasis in the cell if a negative feedback by anthranilate synthase (AS) in tryptophan biosynthesis was altered by gamma irradiation, consequently causing tryptophan over-accumulation. Therefore, the correlation between traits of over-accumulated tryptophan and the alteration of AS by mutagenesis including single-nucleotide polymorphisms (SNPs) would enable the visualization and selection of over-accumulated mutant lines from the rice mutant population. Targeting Induced Local Lesions IN Genome analysis with five fluorescent-labelled primer sets covering the OASA1 locus resulted in the selection of three rice mutant lines, MRVII-9, MRVII-33 and MRVII-37. The tryptophan contents in the seed storage protein of the three mutant lines were elevated when compared with the wild-type cv. Dongan. Genomic DNA of OASA1 was re-sequenced and validated at the SNP and other polymorphism sites. Three mutant lines (MRVII-9, MRVII-33 and MRVII-37) not only contained more than five SNP sites in the total regions of OASA1 gene, but also contained SNPs in coding regions that resulted in amino acid changes such as E241G, A560G and V495F/S496F, respectively. The SNPs, which resulted in amino acid changes, may affect the enhanced tryptophan content in the seed storage protein. These different features could utilize the selection of mutant lines with over-accumulated tryptophan from large rice mutant populations.