Identification of a Seed Phospholipase D Null Allele in Rice (Oryza sativa L.) and Development of SNP Markers for Phospholipase D Deficiency

Abstract

Rice (Oryza sativa L.) bran usage is severely limited by the rapid degradation of rice bran oil and rapid development of hydrolytic rancidity. Because phospholipase D (PLD) disintegrates phospholipid membranes of oil bodies to deteriorate bran quality, we isolated a PLD‐null rice mutant, ‘03‐s108’, whose PLD‐null phenotype is inherited as a single recessive trait. To develop a convenient DNA marker‐assisted selection method for the PLD‐null trait, we first performed a linkage analysis between the PLD‐null phenotype and simple sequence repeat (SSR) markers by using F2 progeny from 03‐s108 × ‘Koshihikari’ crosses, and we mapped the PLD‐null genotype to chromosome 1 near a putative PLD gene sequence, PLDα1. DNA sequencing of the 03‐s108 allele of PLDα1 revealed a transition mutation (G to A) in the third exon. This change resulted in a stop codon, indicating the presence of a nonsense mutation in the 03‐s108 allele. Using this DNA sequence information, we then developed methods of cleaved amplified polymorphic sequence (CAPS) and dot‐blot single‐nucleotide polymorphism (SNP) detection for this mutation, and we confirmed that the sequence polymorphism was predictive of the presence or absence of the seed PLD‐null phenotype. These CAPS and dot‐blot‐SNP methods are easy and cost‐effective means for identifying seed PLD‐null strains, to enable screening and breeding of PLD‐null rice varieties.