Br-related Genes In The Tomato Genetic Transformation Research

Brassinosteroid (BRs) are a class of steroid hormones that play important roles in plant growth and development. BR can promote plant growth and enhance the resistance of plants to environmental stresses. The picture of BR signalling is emerging in recent years. The main components of BR signal transduction include the cell-suface receptor kinase BRI1, the glycogen synthase kinase-3-like kinase BIN2, nuclear proteins BZR1 and BZR2/BES1. A recent study defines BZR1 as a central regulator for coordinating BR signalling, BR biosynthesis, and growth responses.In this study, we constructed an efficient tomato genetic transformation system, and transformed BR signal transduction gene (BZR1 gene) into tomato, which we hope to make a strong basis for further investigating the function of BZR1 in regulating BR signalling, the level of BR in plant, and expression of BR-induced genes, and therefore developing strategy for crop improvement by regulation of BZR1. The main results are as follow:(1) We have developed an efficient method for plant regeneration from tomato explant of cotyledon and hypocotyl. MS medium, supplemented 3% sucrose, 0.7% agar, 2mg/L 6-BA and 0.2mg/L IAA was the optimum medium in obtaining high frequency shoot regeneration in tomato. The frequency of shoot regeneration from cotyledon was 89%, while the frequency of shoot regeneration from hypocotyls reached as high as 94% by this means.(2) The plant expressing plasmid vectors 35S-BZR1-CFP and 35S-bzrl-CFP, which contain BZR1 and bzrl gene respectively were constructed, and introduced to Agrobacterium tumefaciens strain LBA4404 by triparental cross.(3) An efficient gene transformation system was established, and the optimum conditions for the transformation of BZR1 or bzrl were as follow: The explants were infected by Agrobacterium tumefaciens strain LBA4404 for 15 minutes, and cocultivated for 3 days after they were precultivated for 2-3 days, Then the explants were inoculated into the regeneration medium containing 20mg/L Kan to obtain the KanR shoot, and the explants were transferred to fresh medium at intervals of 2-3 weeks. The KanR shoots were cut off from the explants and placed in root-inducing medium when they grew to 2-3cm long. Roots were induced in the shoots after 20 days, and then the cover of the container were opened gradually within the first week, after that, the young transgenic plants were transferred to soil in larger pots, and grown in a greenhouse.(4) 19 KanR young plants were obtained in the tomato genetic transformation test. PCR analysis showed that 16 of the 19 plants contain the 672bp (NPT â…¡) amplification products.