Physiological Characteristics Of A Xanthic Mutant And Fine-mapping Of Its Related Gene Xnt7in Rice

Plants could convert the solar energy into available chemical energy and produce oxygen and organics by photosynthesis. Leaf is the main organ to conduct photosynthesis and95%yields of the rice depends on its photosynthesis, so leaf development will directly affect the rice production. Chloroplasts are the sites for photosynthesis and closely related to the carbon assimilation, whose morphology, structure and numbers directly affect the photosynthesis. At the same time, chlorophyll contents influence the photosynthetic rate too. Chloroplast development and chlorophyll formation are controlled by the nuclear and chloroplast genes whose mutations may lead the chloroplast defected, chlorophyll synthesis damaged and photosynthetic functions frustrated. Rice leaf color mutants are ideal materials to study the photosynthetic system structures and functions, the chlorophyll biosynthesis and the development of chloroplast. A rice leaf color mutant named xnt7was obtained from a japonica variety, Zhonghua11, and died in3leaf stage without chloroplast. Compared with the wild type (Zhonghua11), we studied the morphological characters, genetic analysis, photosynthetic pigment contents and the chloroplast ultra-microstructure in mutants. Meanwhile the XNT7were analyzed and located by mapping, cloning and other molecular biology techniques. The main results were as follows:1The morphology analysis of the xnt7Compared with the wild type Zhonghua11, the mutant sheath was green, while the rest parts were xantha from germination to3leaf stage, the mutant xnt7was died after3leaf (about a month).2The ultrastructure analysis of the xnt7chloroplastCompared with the wild type, The sheath in the mutant xnt7had strange chloroplasts whose internal structures were abnormal and had no obvious matrix and grana thylakoids differentiated. There were a few long and thickness lamella structures while there were no substrate layer structures. The number of granas at other parts of the leaf was less than the wide type and the chloroplast lamella structures dispersed disorderly. Although existing few grana thylakoids in each chloroplast, there were no matrix thylakoids in stroma to link the grana thylakoids, so the mutant couldn’t form a complete photosynthetic membrane system.3Photosynthetic characteristics analysis of the xnt7The contents of chlorophyll a, chlorophyll b, total chlorophyll and carotenoid in mutant sheath were significantly less than that in the wild type but far above the rest parts of the mutant. That illustrated the reason why there were significant differences in color between the sheath and other parts in the mutant.The contents of chlorophyll a, chlorophyll b, total chlorophyll and carotenoid in mutant leaf is significantly less than that in the wild type (Zhonghual1); Due to the total chlorophyll contents decreased largely than that of the carotenoids, mutant xnt7showed xantha.The mutant xnt7had a negative net photosynthetic rate which showed that the mutant conducted respiration only and had no photosynthesis. The mutant couldn’t autotrophic and died.4Genetic analysis of the XNT7The combination of Jinghuil crossed with the xnt7was used to build F1group. The leaf color of F1progenies represented normal green. Then we harvested F2seeds. According to descendant leaf color separation, we built F2group to analyze the gene.In the F2progenies, the segregation ratio of normal green blades(338) to yellow blades(126) was in line with the ratio3:1(x2=1.14<x20.05=3.84), which suggested that the xanthic trait of xnt7was controlled by one pair of recessive nuclear gene.5Fine-mapping of XNT7and determination of its candidateXNT7has been mapped to a region of50Kb between molecular markers R3634-10and R4329-4on chromosome6. XNT7was a new gene because there were no xanthic genes reported in this area. Studied the Rice eFP Browser website, we found eight genes in this region and determined three genes which had strong expression in blade. Then we sequenced the candidate genes. Results showed that LOC_Os06g49670genes lost a G in mutant and the change was in the first exon. LOC_Os06g49670encoded a triangle five peptide (pentatricopeptide repeat, PPR) protein, which composed by repeating35amino acids. In recent years, research has shown that PPR gene exists broadly in plants and there are about650PPR genes in rice; Many leaf color mutants are decided by PPR abnormal synthesis. Combined above results, we finally determined the gene as XNT7candidate.