| Rice (Oryza saliva L.) is an important food crop, enhance its production and improve its quality is the primary task of rice breeding. Rice yield is constituted spike per plant, grains per spike, grain weight. Grain size directly affect grain weight and thus determine rice production, therefore, the mapping and cloning related genes of grain size contributes to depth understanding of the mechanism of grain formation and provides a theoretical basis for improving yield. In this study, a mutant Nanjing35 from 60Co-y radiation mutant library appeares dwarf, delaying heading, erect leaves, short and compact spike, smaller grain, significantly decreased grain weigh, etc. characteristics. Grain filling rate of Nanjing35 and mutants was measured and we found that grain filling rate of mutant was lower than the wildtype and dry matter accumulation was slower. Secondly, we observed that smaller epidermal cells and a higher cell density in the outer surface of glumes in mutant with ascanning electron microscopy, while allowed a smaller glumes and thus resulted in slow filling and a smaller grain. F1 and F2 obtained from mutant and Nanjing35 hybridization were similar phenotypic to Nanjing35 and phenotypic separation, respectively. F2 population appeared normal plant height and grain seeds and dwarf smaller grain and the ratio of 3:1, indicating that the grain mutant gene is controlled by a single recessive gene. In order to isolate the mutant gene, we selected extreme individuals from F2 segregating population from the mutant and Dianjingyou hybridization, and utilized marks evenly distributed on 12 chromosomes existing laboratory and analysed genotype of extreme individuals. First 48 extreme individuals from 500 groups were used to linkage analysis, and we initially located mutant loci on the long arm of chromosome 5 within 2434.8kb interregion. Then by expanding mapping population to 5900 and picking out 620 extreme individuals and developing polymorphic markers, we positioned the mutant gene into the 230kb region eventually. Using the Gramene database for prediction genes, we found 23 genes in location section and 15 genes with functional annotation, which established foundation for map-based cloning of mutated gene.WRKY family genes is a class of transcription factor contained more members in rice (Oryza sativa L.), it is not only in response to biotic and abiotic stress, but also involve plant morphogenesis. WRKY transcription factors include WRKYGQK motif and zinc finger domain and WRKY family genes can be divided into four subgroup according to the number and type of WRKYGQK motif and zinc finger domain in rice. In this study, a mutant T387 was screened from the T-DNA insertion mutant library in rice, which appeared semi-dwarf plant, smaller brown rice and significantly decreased grain weigh. The insertion site of mutant T387 was predicted through the website (OryGenesDB a database for rice reverse genetics), which was verified by two pairs of primers. T-DNA insertion located in the gene the 5’-UTR region of OsWRKY36 does not affect OsWRKY36 transcription in the T387. Because of the presence of enhancer label of T-DNA, the expression of OsWRKY36 significantly upregulated in the T387. By analyzing OsWRKY36 proteins, TaWRKY3 and ZmWRKY36 were highest homology with OsWRKY36. To understand the biological function of OsWRKY36, overexpression and RNAi and PooswRKY36:GUS vector were constructed, which verified expression model and controlled phenotype of OsWRKY36 by transgenic plants. |
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