关键词: 水稻 , 斑马叶突变体 , 遗传分析 , 类胡萝卜素 , &beta, -OsLCY

Abstract: 【Objective】 The present study was aimed at morphological characterization and candidate gene analysis of zebra leaf mutant zebra524 in rice, so as to lay the foundation for further functional analysis of this gene and its application in agricultural production. 【Method】 A zebra leaf mutant, designated as zebra524, was isolated from an ethyl methanesulfonate (EMS)-mutagenized population. Morphological characteristics and the agronomic traits of the mutant and its wildtype were contrasted systematically. Using spectrophotometer, photosynthetic pigment contents in leaves of the mutant and its wildtype were measured at seedling and booting stages, and in glumes at grain-filling stage, respectively. After the zebra524 mutant was crossed with normal green varieties, the leaf colour phenotypes of the F1 progenies and the segregation ratio of zebra leaf plants to green leaf plants in the F2 populations were investigated. A total of 575 zebra leaf mutant individuals in the F2 mapping population generated from the cross between the mutant and G46B (Indica) were used for mapping using molecular markers. In addition, putative genes in the mapped region were analyzed, and candidate genes in the mutant and its wild-type were sequenced, respectively. Subsequently, DNA sequencing of β-OsLCY gene and alignment of the deduced amino acid sequences of homologous β-OsLCY proteins were conducted. 【Result】 During seedling and tillering stages, the zebra524 mutant showed alternating transverse pale green and albino sectors on leaves. During booting stage, its albino sectors turned into pale green and hereafter, the newly grown leaves became completely pale green. The zebra524 mutant also showed pale glumes during heading and grain-filling stages, and brown nodes, pink basal internodes, orange-red embryos and basal caryopsises at maturation stage. In addition, plumules of zebra524 showed an orange-red color in the dark. Chlorophyll contents in leaves of the zebra524 mutant decreased by 82.8% and 20.9%, and carotenoid contents decreased by 64.7% and 32.6% at seedling and booting stages, respectively, compared with those of the wild type. In addition, the levels of chlorophylls and carotenoids in glumes of zebra524 mutant were reduced by 38.1% and 42.8%, respectively, at the grain-filling stage. Its plant height, number of spikelets per panicle, seed setting rate and 1000-grain weight were reduced by 12.3%, 9.5%, 13.0% and 5.4%, respectively, at maturation. All F1 plants displayed wild-type phenotype, and the number of normal seedlings versus that of mutant seedlings in the F2 populations was fitted to 3:1, indicating that the zebra524 phenotype was controlled by a single recessive nuclear gene. The mutant gene was mapped to a region of 235 kb between SSR marker RM7082 and InDel marker Y2 on the short arm of chromosome 2. Sequencing analysis of candidate genes between the mutant and its wild-type revealed a single-nucleotide G-to-T mutation was found at position 235 in the coding region of the LOC_Os02g09750 gene for lycopene β-cyclase, which resulted in an amino acid change from Gly at position 79 to Cys in the encoded product. 【Conclusion】 The zebra524 mutant gene was allelic to β-OsLCY gene which was documented previously. The zebra524 mutant phenotype may be attributed to a point mutation of β-OsLCY gene encoding lycopene β-cyclase.

Key words: rice , zebra leaf mutant , genetic analysis , carotenoid , β-OsLCY gene