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Optimizing the Techno-system of Genetic Transformation Mediated by Agrobacterium Tumefaciens and LeJA2 Gene Function Research of Tomato

Author: LiuZuoZuo
Tutor: ZhangChunQing
School: Shandong Agricultural University
Course: Crop Genetics and Breeding
Keywords: Tomato Tissue culture Genetic transformation Jasmonic acid Signal Transduction Systemic resistance response
CLC: S641.2
Type: Master's thesis
Year: 2009
Downloads: 147
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Abstract


At present, the application of large doses of pesticides and herbicides on the environment caused a lot of damage, also caused a bad effect on the human body and quality of life. Higher plants in the long-term evolution of complex and sophisticated resistant response mechanism to resist against insects. In-depth understanding of plants to insect resistance molecular mechanism will provide a theoretical basis for the use of the plant's own resistance to the establishment of environmentally friendly strategies to control agricultural pests. The identification of signaling molecules including the polypeptide signal the molecular system hormone (Systemin) and unsaturated fatty acids derived from the plant hormone jasmonic acid (JA), both mediated resistance reaction from the molecular level of genetic resolution, identified one of the important components (functional genomics), and features in-depth study. This study to optimize the tomato genetic transformation system; while taking advantage of the principle of reverse genetics, built LeJA2 (Lycopersicon the es-culentum jasmonic acid 2) overexpression vector and RNAi expression vector, Agrobacterium-mediated transgenic technology, access to two types of transgenic seedlings, determination of character identification and PPO activity of the two types of transgenic seedlings, LeJA2 function to preliminary estimates. The main results of the study are as follows: (1) to establish a system of Agrobacterium-mediated genetic transformation of tomato tomato conversion LeJA2 of Agrobacterium-mediated gene system has been optimized Agrobacterium infection, infection temperature and infection Concentration factors of tomato callus formation, to seek a major factor affecting the genetic transformation, to obtain a better tomato Agrobacterium infection. 2 transgenic tomato plants transformed cotyledons receptors, the use of efficient Agrobacterium-mediated genetic transformation system the LeJA2 gene and LeJA2-Rani gene into tomato, 20 LeJA2 overexpression of tomato resistant plants and 35 LeJA2 gene silencing resistant plants, PCR test results showed that the kanamycin-resistant strains can amplify the target band of 1050 and 517bp, to prove that the gene was integrated into the tomato genome. Functional verification PPO activity in transgenic tomato plants were detected. On PPO activity test: after the plant mechanical damage before, LeJA2 gene overexpression transgenic plants PPO dyeing dark brown, still showing control and turn LeJA2-RNAi transgenic plants PPO staining the normal juice of green. With scissors after mechanical injury PPO the staining, LeJA2 gene overexpression plants turn LeJA2-Rani transgenic plants and control staining dark brown. The results showed that the gene LeJA2 injury tomato jasmonic acid-mediated signal transduction pathway plays an upstream regulatory role in regulating expression of the resistance gene.

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CLC: > Agricultural Sciences > Gardening > Vegetable gardening > Solanaceous > Tomatoes ( tomatoes )
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