Isolation And Characterization Of Salicylic Acid Induced Transcription Factor SlWRKY From Tomato

Tomato is an important economic crop, but it faces serious declines in yield and quality as a result of attack by various environment stress. Nowadays, it has been a serious problem. Studies have shown that WRKY transcription factors can bind to W-box of the promoter of stress resistance genes in plant biotic and abiotic stresses resistance, so the WRKY transcription factor has become a hot spot for crop improvement. Currently, the study of tomato WRKY transcription factor focused on cloning gene fragments, rare of function analysis. In this study, we cloned two tomato WRKY genes and analysed their functions. These would contribute to reveal the function of tomato WRKY transcription factor and breed obtaining resistance plants.Previous studies have found that SA can induce the expression of WRKY transcription factors, however, the concentration and the best way of SA treatment have not been reported. Two treatment methods and four concentration of SA were designed, then analysed defensive physiological parameters of tomato. The results showed that root treatment was slightly better than the leaves spraying; In 0-2mM range, POD, PAL, PPO and protein content of tomato leaf increased with the SA concentration, especially in the the concentration of 2mM they reach the higest level, which can be used to clone resistance WRKY transcription factor.In this study, we obtained two full-length of tomato WRKY transcription factor genes, designed SIWRKY1 and SIWRKY2. SIWRKY1 length of 974bp, including 762bp coding region, 114bp 5'UTR,98bp 3'UTR; SIWRKY2 length of 2152bp, including 1659bp coding region, 130bp 5'UTR, and 363bp 3'UTR. Semi-quantitative analysis showed that,400mM NaCl or 4℃could induce SlWRKY1 and SlWRKY2 expression; ABA and SA can induce the expression of SIWRKY2; the expression of SIWRKY2 could be detected in multiple tissues and organs, but the level of mature tissue is much higher than immature organizations; the highest expression in flowers, followed by seeds.To elucidate the function, open reading frame of SIWRKY2 was introduced into tobacco, and the expression of SIWRKY2 was detected in transgenic tobacco plants by PCR and RT-PCR analysis, respectively. The results demonstrated that overexpression of SIWRKY2 in tobacco plants enhanced tolerance to salt and drought stress. Semi-quantitative PCR results showed that, the expression of PR1 and PR2 in transgenic tobacco were significantly increased, indicating S1WRKY2 can interacte with PR1 and PR2 then played a regulatory role in improving the plant's resistance.In order to further reveal the mechanism of stress resistance of transgenic tobacco, the chlorophyll content, POD, SOD activity, the relative conductivity and MDA content were measured after salt and drought stress treatment. The results showed that with the stress intensity increased, the relative conductivity and MDA content of transgenic tobacco lower than the wild type; chlorophyll content relatively stable, while the wild type were significantly lower; SOD and POD activities increased faster than the wild type. Among the different transgenic lines, Line 17 has best ability to resistance stress.