Genetic Analysis, High-resolution Mapping And Physiological Characteristic Of A Premature Senescence Mutant Pse(t) In Rice (Oryza Sativa L.)

The Zhonghua 11 (Oryza sativa L. subsp.japonica) was used for transformations to construct the T-DNA inserted poll with the method mediated by Agrobacterium tumefaciens strain containing the binary vector pDsBar1300. The line T2432 was isolated from the T₁ generation in the T-DNA inserted poll, the premature senescence symptoms appeared more seriously in 13 mutants compared with the control plants at the late development stage. The premature senescence mutant is not reported so far, the mutant was named pse(t) (premature senescence, tentatively) based on the similar mutant in maize. The pse(t) mutant provided a good material to study the molecular mechanism underlying the premature senescence at the late development stage of crops. The physiological characteristics of pse(t), genetic mechanism and molecular mapping was carried out in this paper.In two phytotrons under controlled at the 24 ℃ and 28 ℃, the content of chlorophyll, soluble protein and MDA, net photosynthesis, the antioxidant enzymes activities of SOD (EC 1.15.1.1) and POD (EC 1.11.1.7) and the activities of the peptidase of leaves in pse(t) were measured from top to bottom according to the leaf positions at the flowering stage. Compared with the control plant, the mutant showed the characteristics: 1, higher net photosynthesis rate (Pn) appeared in the 1st and 2nd leaves, the contents of chlorophyll and the soluble protein were also higher in the 1st leaf. 2, the symptom of premature senescence was accelerated in mutant in the high phytotron (28 ℃). 3, the trends of the activities of SOD, POD and peptidase were higher in the 3rd and 4th leaves. Under the two temperature treatments, the content of MDA, the activities of SOD were no significant difference at the same leaf position of mutant, and the activities of POD as well. The content of chlorophyll and soluble protein of leaves in mutant decreased rapidly in the 28 ℃ treatment. These results showed that the pse(t) was sensitive to high temperature.Genetic analysis of the pse(t) mutant revealed that the phenomenon of premature senescence was controlled by a single recessive nuclear gene. Further studying resultindicated that the mutation was not induced by T-DNA insertion. In order to improve our understanding of the molecular mechanism for premature senescence in rice at the genetic and molecular levels, a map-based cloning strategy was employed to locate the Pse(t) gene. The indica-japonica F2 population was constructed by the conventional crossing method (pse(t) X Longtepu(zW/ca)). The newly development SSR and CAPS markers were designed based on the rice genome sequence(http://www.ncbi.nlm.nih.gov/BLAST/Genome/PlantBlast.shtml77) ,46 pairs new SSR and CAPS markers were developed, 58 percent markers appeared the polymorphism between the Zhonghua 11 and Longtepu. Results of the genetic linkage analysis showed that the Pse(t) was located in the genetic distance of 3cM between the markers RM418 and R646 on the long arm of chromosome 7. A high-resolution mapping of the Pse(t) locus was carried out using SSR and CAPS markers by the F2 populations comprising of 1,961 individual mutants and 2,002 wild type plants. The Pse(t) locus was found to be flanked by markers SS22 and PP21 at 0.06 cM and 0.09 cM intervals, respectively, with the corresponding distance of 220-kb; the Pse(t) locus was found to co-segregate with three markers (SSI5, PP4 and SS20). These findings provide the important information for the final isolation of the Pse(t) gene.Gene prediction indicated that there were 10 full-length cDNAs in the region of 220-kb region (http://ricegaas.dna.affrc.RO.ip/rgadb/). Based on the physiological analysis, there are two candidate genes in the region. One of the candidate genes participated in the programmed cell death and protecting functions under high temperature conditions; the other candidate genes presumably takes part in the electronic transport process associating with the free radical accumulation. The further cloning the Pse(t) and function complementation were in progress.