Characterization And Gene Mapping Of Two Leaf Colour Mutants In Rice

Leaf colour mutant is an useful resource for rice genomics research. In this study, we found twoleaf colour mutants in rice: green-revertible albino mutant and lesion mimic mutant. We conductedmorphological identification, genetic analysis, determination of chlorophyll content and electronmicroscopy observing of chloroplast development of the two mutants. The two mutants were finemapping by map-based clone method. Those results were helpful to illustrate the molecularmechanism of leaf colour mutant. The results are summarized as follows:1) A stable genetic green-revertible albino mutant was obtained from the progeny of a Japonicavariety Zhonghua11treated with EMS irradiation. Under the different temperature, the mutantdisplayed distinctive albino leaf at the first leaf stage, displayed lighter white stripes at the secondand third leaf stage, at the forth leaf period the mutant begun to turn green gradually. Temporarydesignated as green-revertible albino mutant1（gra1）. Compared with wild type, the gra1mutantplant height, seed setting ratio and1000-grain weight were significantly reduced. Results ofelectron microscopic showed that the white part of the gra1third leaf displayed less mesophyllcells and the chloroplast microstructure showed abnormal compared with wild type, the green partof the gra1third leaf showed normal chloroplast structure. By the map-based cloning, the targetgene was finally narrowed down to a230.3kb region between Indel10and Indel20on the long armof chromosome1. The sequencing analysis results revealed that the21st ORF (Os01t0851000-02),annotated Carbohydrate/purine kinase domain containing protein, carries a single nucleotidemutation (G to A) compared wild type sequence, the result implied that the Os01t0851000-02might be the candidate gene of gra1. RT-PCR assay showed that chlorophyll biosynthesis genesPORA, chloroplast development-related genes V1, photosynthesis-related genes psaA, psbA andrbcL showed significantly up regulated and the only chlorophyll biosynthesis genes YGL1showedsignificant down regulated compared with wild type. The results showed that gra1play animportant role in rice chlorophyll synthesis and chloroplast development.2) The lesion mimic mutant lmm4was selected from the mutant bank of Nipponbare treatedwith EMS, brown lesions were first observed on the tip of lmm4leaves at initial Tillering stageand spread gradually downward to the whole leaf blade and even the leaf sheath. Compared withwild type, the chlorophyll and the net photosynthetic of lmm4were significantly reduced. Theplant height, panicle length, seed setting rate, grain weight also decreased significantly. Theshading assay show that the lesions on lmm4leaves were induced by natural light. The stain assayshow the lesions on lmm4leaves were caused by the cell death and the lmm4leaves hadaccumulation of hydrogen peroxide. Inoculated assay show that lmm4displayed significantresistant to blast race C7and G1compared with wild type. Pathogenesis-related gene expressionanalysis show that POC1、PAL、PBZ1、PR1were high express compared with wild type. Geneticanalysis showed that the phenotype of lmm4was controlled by a single recessive nuclear gene. Bymap-based clone method the lmm4finally narrowed down to a235kb region between Indel-a4and Indel-a6near the centromere of chromosome5.