Rice leaf senescence control gene ES2 and application thereof
A leaf senescence and rice technology, applied in the field of rice leaf senescence control gene ES2 and its application in rice breeding, can solve the problems of leaf stomata shrinkage, photosynthetic ability decline, cell proliferation and differentiation ability and physiological function decline, etc.
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Embodiment 1
[0046] Embodiment 1, the acquisition of mutant material
[0047] Through chemical mutagenesis of japonica rice variety Taipei 309 by EMS, a premature leaf senescence mutant es2 was screened. The traits of the mutant have been stably inherited after multiple generations of selfing, and the premature senescence phenotype begins to appear at the beginning of tillering, and its growth and development are obviously slower than that of the wild type, and the tillering is obviously less than that of the wild type, and the premature senescence phenotype of the leaves is very obvious at the heading stage . All rice materials were planted in the experimental field of School of Chemistry, Zhejiang Normal University, Jinhua City, Zhejiang Province, under conventional management.
[0048] Phenotypic analysis of wild-type Taipei 309 and mutant es2 as in figure 1 shown.
[0049] The above-mentioned EMS chemical mutagenesis method is as follows: immerse the seeds of Taipei 309 in ethyl met...
Embodiment 2
[0050] Embodiment 2, population construction and genetic analysis
[0051] The mutant es2 was crossed with conventional indica rice TN1 and ZF802, and F 1 The plants showed normal wild-type phenotype, indicating that es2 was controlled by a recessive nuclear gene. Statistics F 2 Segregation population segregation ratio (Table 1), the results show that the segregation ratio of the plants of normal phenotype and mutant phenotype is close to 3:1 segregation through Chi-square test, which shows that the leaf premature senescence phenotype of es2 is caused by single recessive Nuclear gene control.
[0052] Table 1 Genetic analysis of leaf premature senescence mutant es2
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Embodiment 3
[0055] Embodiment 3, fine mapping of ES2 gene
[0056] The mutants and TN1 were screened for polymorphism by using the SSR primers evenly distributed on 12 rice chromosomes preserved in our laboratory, and 116 pairs of SSR primers were screened for polymorphism. Then use 21 F in es2 / TN1 2 Linkage analysis was performed on individual plants with premature senescence to preliminarily confirm the chromosomal location of the target gene. Genomic DNA was extracted by CTAB method. Specific steps are as follows:
[0057] ①. Weigh 0.1g of rice leaves and grind them into powder with liquid nitrogen, then add 600 μl of CTAB solution (2% (m / V) CTAB, 100mmol / L Tris-Cl, 20mmol / L EDTA, 1.4mol / L NaCl ; pH8.0) prepared DNA extraction buffer, 65 ° C water bath for 40 minutes. Add another 600 μl of chloroform:isoamyl alcohol (volume ratio of 24:1), and mix well. Centrifuge at 10,000rpm for 5 minutes and transfer the supernatant to a new centrifuge tube.
[0058] ②. Add 2 / 3 to 1 volume of ...
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