Characterization And Gene Cloning Of A Yellow Green Leaf Mutant In Rice

Leaf colour mutant is an important resource for Photosynthesis research. A yellow green leaf mutant was isolated from rice tissue culture transgenic lines of rice(Oryza sativa L. subsp. japonica cv. Nipponbare). In this study, this mutant was used to conduct research on phenotypic indentification, genetic analysis, gene clone and functional analysis.The results showed as follows:The mutant has showed yellow green leaf since the seedling stage, so it was named ygl12. Analysis of chlorophyll content showed that the chlorophyll content of mutant decreased Compared to wild type, and the chlorophyll a reduced to a great extent. Temperature/light intensity sensitive test indicated that the mutant phenotype was insensitive to temperature, but it was sensitive to light intensity. The chloroplast microstructure showed abnormal development in the mutant. Compared with wild type, the mutant flowering time was delayed, the mutant productive tiller number, seed setting ratio and 1000-grain weight reduced, but the flag was larger, the main Panicle was longer.Genetic analysis revealed that the mutation is controlled by a single recessive gene. A hygromycin resistance assay showed that the mutation was not caused by T‐DNA insertion, so a map-based cloning strategy was chosen to isolate the ygl12. A cross between the ygl12 mutant and Longtepu composed the Mapping population, using 21 F2 mutants, the ygl12 gene was roughly mapped into 6.5c M between SSR marker ML7 and L55 on the chromosome 12. A high-resolution physical map of the chromosomal region around the ygl12 gene was made using F2 population consisting of 1776 mutant individuals. Finally, the ygl12 was mapped within a 76.5kb interval between two INDELS markers YG4 and YG5. There were 12 ORFs in the targeted region by querying The Rice Annotation Project Database, Sequencing results further revealed that there is 39 bp deletion in LOC_Os12g23180, therefore, LOC_Os06g10350 may be the target gene, Transgenic complementation rescued the ygl12 phenotype verified LOC_Os12g23180 was the YGL12 gene.RTFQ PCR assay showed that most chlorophyll biosynthesis genes expression increased suggested that YGL12 may play an important role in chlorophyll synthesis.The YGL12 gene transcript analysis in different organs indicated that it was mainly expressed in green organs especially in leaf. Subcellular localization experiment show target protein was localized in chloroplast.CSP41 b that is Chloroplast stem–loop-binding protein that binds RNA in Arabidopsis thaliana is the homologous protein of YGL12 that suggested YGL12 may work as the same as CSP41 b. These results will be helpful for dissecting the function of YGL12 gene.