Genetic Analysis And Gene Mapping Of A Novel Rolled Leaf Mutant Rl12(t) In Rice

Rice (Oryza sativa L.) is one of the most important cereal crops in the world. With the increasing demands for food and continuing declines of the cultivated lands, how to improve the yield of rice has become the most important issue for breeders and cultivation exporters. In recent years, the "ideal plant type" models for high-yielding rice are quite a lot, and some them referred to the straight and roll of rice leaves. Rice leaves are vital organs for photosynthesis, and recent research also showed that rice leaves play an important role in high yield breeding. Its moderate rolling could improve plant population's structures and enhance the light-use efficiency. The rolled leaf mutants are significant resources in ideal type breeding. It was probably that moderate rolled leaf has built a good platform for plants to receive as much light as possible. Meanwhile, rolled leaf is often accompanied by the improvement of chlorophyll content, which is an important component for the light absorption and transformation. Thus, rolled leaf resources have important utilizable value in high photosynthetic efficiency breeding.There are plenty of rolled leaf resources with genetic stability in rice, but genetic studies on these rolled leaf genes are still lacking. Currently, including six genes been mapped by the classical genetic methods,17 rolled leaf genes had been positioned with 15 recessive genes, one incompletely dominant gene and one incompletely recessive gene. Until now, no completely dominant rolled leaf gene has been mapped. In this study, we report a novel rolled leaf mutants, rl12(t), derived from the restorer line Jinhui10 (Oryza sativa L. ssp. indica) treated by EMS. This mutant trait, varied during the growth period, was controlled by a dominant nuclear gene. The morphological analysis, physiological analysis, cytological analysis, agronomical characters analysis, genetic analysis and gene mapping of this rolled leaf mutation was reported in this study. The results as follows:1. Morphological AnalysisCompletely different from the leaf shape previously reported, leaf curling characteristic of rl12(t) presented from tillering stage and been changing during the growth process. Leaf curled inward along the veins. The main performance is that the new leaves didn't roll, however, the mature leaves curled the upper 1/3 section of them and the older mature leaves rolled completely. When the flag leaf out, it also showed rolled leaf trait with the upper 1/3 leaf curling.2. Physiological Analysisleaf pigment content of rl12(t) and wild-type was extracted at tillering, jointing, heading and maturity stage. The results showed that the chlorophyll content of rl12(t) was higher than that of the control in any period. Especially in the heading stage, the contents of chlorophyll a, chlorophyll b, total chlorophyll and carotenoids of rl12(t), were significantly higher than that of the control Jinhui 10. Therefore the rolled leaf mutant increased the pigment contents in different periods, provided a good foundation for the improvement of the photosynthetic rate of rice.3. Cytological AnalysisIn order to find the main reason responsible to the formation of rolled leaf, we took the fresh leaves of the rl12(t) mutant and the control in the period of the rolled leaf trait fully performanceexpressed, and made them into paraffin sections. Cell structures observed under the electron microscope showed that, in the rolled leaf mutant, mesophyll cells had great distributions under the leaf epidermis and scattered all over of the small vascular bundles of leaf. In the control, mesophyll cells in both sides of the leaf cells were well-distributed. Contrary to the mutant, hardly any mesophyll cells distributed along the small vascular bundles of the leaf. These differences may be due to the different curvature of the movement cells between the small vascular.4. Agronomical Characters Analysisrl12(t) rolled leaf mutant demonstrated rolled leaf traits in the tillering stage. During the whole period of the vegetative growth, other characters such as plant height, leaf angle, tiller number and normal plants had no significant differences, besides the ABSTRACT leaf curl. While, after the period of mature and heading, there are no significant differences in panicles, panicle length and total grains between rl12(t) mutant and wild-type (Jinhui 10). However, the seed setting rate, panicle of the mutant was significantly lower than that of the wild type, which showed that this rolled leaf gene may also affect the setting rate, grain weight, panicle, and other important agronomic traits.5. Genetic Analysis of the rolled leaf traitIn this study, we hybrid the rolled leaf mutant rl12(t) and the flat leaves of the CMS parent Xinong 1A. F1 leafs showed exactly the same phenotype as the rl12(t), which indicated that the mutant is controlled by a dominant gene. There is a clear separation in F2 populations, which performance characters of both parents, without the middle type of leaf emergence plants. Curly trees (4160):normal trees (1345) full compliance with 3:1 segregation ratio (x2= 0.9161<3.84 [x2 (0.05,1)]), which showed that the leaf mutant is controlled by a pair of completely dominant single gene.6. Molecular Mapping of the Rolled Leaf Gene400 SSR markers which uniformly coverage the whole rice genome were used to screen the wildtype and the mutant gene pools for polymorphisms. Two markers, RM3590 and RM5348, on the chromosome 10 showed polymorphisms between these two gene pools.10 mutant and 10 normal individuals of F2 were used for strains linkage analysis, markers RM3590 and RM5348 are linked with the target traits according to the linkage analysis. Then all of F2 mutant individuals were screened by these two markers and then the target locus was tentatively located between them, with the genetic distances of 12.1 cM and 2.6 cM respectively. In order to further localize RL12(t) gene to a smaller region, new SSR markers were developed between markers RM3590 and RM5348 according to the published SSR markers (http://www.gramene.org/) and genome sequences of Indica cultivar 9311. Finally, RL12(t) gene was located between markers SWU-1 and SWU-2, with the genetic distances of 1.8 cM and 0.6 cM respectively.