Genetic Analysis And Gene Mapping For A Yellow-green Leaf Mutant Ygl11(t) In Rice

The leaf is the most important organ where photosynthesis ocurres and it plays an importent role in the growth and development process of rice. Characterisation of the rice leaf color mutants provides not only important materials for studying the regulation, synthesis and degradation of chlorophyll as well as the development of chloroplast and the photosynthesis, but also the specific germplasm resources for rice breeding. Thus, the genetic analysis, cloning and functional studies of genes related to leaf color in rice are of great significance in theory and practice.In the present study, a novel leaf mutant isolated from an indica variety 93-11 via radiation of 60Co-y in M2 generation was characterized. We temporarily named this mutant as ygl11(t) (yellow green leaf). In order to investigate the mechanism underlying the mutant phenotype, the morphological, agronomic traits, chlorophyll contents, chloroplast development and fluorescence parameters among the ygl 11 (t) mutant and its wild type were detaildedly evaluated. Also, genetic analysis of the mutant was conducted and the YGL11(t) gene was finely mapped. Then the gene prediction was done within the located region. The main results are as follows:1. The whole plant of mutant displayed yellow-green leaves at the seedling stage, while light green at the heading stage. The mutant plants have normal seed setting, but the growth potential weakened significantly when compared to the wild type. For example, the plant height reduced by 8.1%, effective panicles 37.0%, total grains 18.5%, and grain weight 6.9%.2. The ygl11(t) mutant is quite different from chlorina mutant, and exhibits delayed greening during photomorphogenesis because of slow rates of chlorophyll accumulation. Eventually, the mutant plants accumulated substantial quantities of chlorophyll, similar to the wild-type levels in the matured leaves. From seedling to heading stage, the mutant leaves gradually turn green. Meanwhile, the content of chlorophyll a, chlorophyll b and total chlorophyll in mutant gradually increased from 58.47%,23.77% and 49.9% of wild type to 93.56%,76.56% and 89.8%, respectively. Chlorophyll a/b ratio also dropped to 4.3 from 7.6. These results indicated that the synthesis of photosynthetic pigments was severely suppressed in the seedling stage.3. Transmission electron microscopy analysis showed that, the chloroplast in the mutant ygl11(t) developed slow behind the wild-type. At seedling stage, although the chloroplast structure of mutant was normal, less grana stacks and scattered distribution were observed when compared with the wide type in developing leaves. At the tillering stage, the interval between the layers became larger, the vesicle-like structures were rich and the thylakoid body stamp stack is still small, as a whole, the entire chloroplast still dysplasia. However, at the filling stage, the chloroplast in mutant leaves came close to that of wide type.4. Fluorescence kinetics analysis showed that, the photochemistry in both donor side and acceptor side were significantly inhibited in photosystem II (PSII) of ygl11(t) mutant, also the photosynthetic capacity and light protection ability were reduced. The maximum photochemical efficiency of PSII (Fv/Fm) had no significant difference between ygl111(t) and the wild-type at the seedling stage, indicating that photosynthetic system structure had not been destroyed. However, the actual photosynthetic efficiency of PSII (Y (II)), photochemical quenching (qP), PSII electron transport rate (ETR) and non-photochemical quenching (NPQ) were found to be decreased by 27.7%,27.4%,33.6% and 21.62%, respectively, indicating that both the energy absorption and utilization of light energy in its light systems for each chloroplast in yellow-green leaves were significantly reduced.5. Genetic analysis showed that, the yellow green leaf trait of ygl11(t) mutant was controlled by a single recessive nuclear gene. The gene, named YGL11 (t), was narrowed to the 42kb region on the short arm of chromosome 11, between the Indel markers s11-51 and s11-58.6. There are 9 ORFs in this region, and two of them, LOC_Os11g05552 and LOC_Osllg05556, are associated with chloroplast development, encoding highly homologous signal recognition particle 54kDa protein (cpSRP54). As known, cpSRP54 is related to the translocation of chloroplast membrane protein, so we sequenced these two genes firstly. The results showed that there was no sequence differences in LOCOs 11g05556, but 20 bases were missed in the sixth exon (coding region 637-656bp) of LOCOs11gO5552 in ygl11(t), leading to frame-shift and delayed termination of amino acid translation. An InDel marker D5552 designed based on this 20bp deletion was co-segregate with the YGL11 (t) gene. So we infer that the LOC_Osllg05552 gene is the candidate gene of YGL11 (t). It is a new allele of YGL138 (t).7. The Real-time PCR results of LOC_Os11g05552 and LOC_Osllg05556 displayed a similar tissue-specific expression pattern, mainly expressed in green tissues, with the the highest expression level in leaves, followed by the sheath, young panicles, stems and roots. In addition, the LOC_Os11g05552 did not express in mutant, while LOC_Os11g05556 express patterns were similaramong the ygl11 (t) mutant and its wild-type. The fact that the mutant leaves gradually turn green in its later growth stage, it is infered that LOC_Os11g05556 could compersate the loss-of-function of LOCOs11g05552.