Cloning And Expression Analysis Of LeCTR4svZS And Obtaining Of Transgenic Plants

Ethylene signal transduction negative regulatory factor RAF protein kinase CTR in plant growth plays an important role in the development process, the tomato CTR belong to a multigene family:LeCTR1,LeCTR2,LeCTR3,LeCTR4 members, which LeCTR4 have multiple shear the, its biological role is still unclear. In the condition of ethylene, the expression of LeCTRl was increased, with the maturity of the fruits, the expression of LeCTRl increases. LeCTR3 and LeCTR4 high expression in the leaves, and is not sensitive to ethylene. It is indicated that the members of multi gene family have different functions. The study by cloning LeCTR4svl found a new LeCTR4svZS transcripts and analysis found its RAF like protein kinase CTR4 structure, and miR1917 splice sites, and by means of tomato tissue specific expression of and to clarify its in tomato development process in the temporal and spatial expression pattern, through the construction of LeCTR4svZS and over expression plants of anti miR1917 shear mLeCTR4svZS. To lay the foundation for further understanding the ethylene signal transduction pathway and alternative splicing. The main results are as follows:1. In the vegetables in six tomato, cloned CTR4 a new transcription of the CTR4svZS, based on sequence alignment, sequence than the NCBI database CTR4sv1 ORF much a sequence of, using bioinformatics tools found CTR4svZS basic characteristics of CTR gene family, recognized to belong to CTR4 the other a shear mode for the CTR4svZS. But its biological function is not clear.2. The tissue specific expression analysis of CTR family genes showed that the expression levels of different CTR genes were different in the growth and development stages of tomato. At the same time, the expression of CTR3, CTR4 and CTR4svZS genes in white mature period increased rapidly, indicating that these CTR genes may be involved in the ripening of fruit. The CTR gene in root, stem and leaf in expression was similar, suggesting that these genes may be involved in the process of growth and development of Tomato in multiple organs and redundancy.3. The expression of CTR1 gene was relatively stable under ethylene condition, which indicated that exogenous ethylene had no effect on the expression of CTR1 gene. The expression of CTR4svZS and CTR4 was regulated by exogenous ethylene. And the expression of ethylene from pedicel analysis are compared, CTR3, CTR4, CTR4svZS expression in response to exogenous ethylene.4. miR1917 and target genes CTR4svZS in ripe fruit and pedicel abscission expression was negatively correlated, suggesting that the fruit ripening and abscission process miR1917 of CTR4svZS negative regulation to an important role the early stage in the process.5. Amplification of full-length sequence of CTR4svZS, and site directed mutagenesis of miR1917 degradation sites, the successful construction of 35S::CTR4svZS and anti miR1917 degradation of 35S::plant mCTR4svZS expression vector, into in vegetables, tomato, and carry out quantitative PCR and real-time screening of transgenic tomato and get the 35S::3 strains of CTR4svZS transgenic plants and 35S::mCTR4svZS transgenic plants 4 strains. Laying a solid foundation for the follow-up functional verification test.