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| Cloning and Expression Analysis of Transcription Factor TwWRKY1 Gene in Tripterygium wilfordii |
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Abstract As a traditional medicine, Tripterygium wilfordii, which the main active ingredients are terpenes and alkaloids, has a long history and wide range of applications in agriculture and medicine. The WRKY transcription factor is a plant-specific transcription factor and involves in plant growth, development, aging, biotic and abiotic stress and secondary metabolites biosynthesis regulation. From the hairy roots of T. wilfordii, This study cloned a WRKY transcription factor and named it as TwWRKY1 (GenBank No. GAVZ01022264.1) using rapid-amplification of cDNA ends (RACE) technology. The full-length of the gene is 962 bp, the open reading frame is 537 bp and encodes 178 amino acids. The analysis of bioinformatics showed TwWRKY1 contains a typical WRKY conserved domain, zinc finger structure was C2H2, which belonged to Class Ⅱ of WRKY family; the phylogenetic analysis showed that the TwWRKY1 has a 83% parental relationship with Coptis jiponica CjWRKY1. Analysis of TwWRKY1 expression in different tissues showed that the gene constitutively expressed in roots, stems and leaves and the expression level was the highest in young leaves and was the least in natural root using qRT-PCR. The expression of TwWRKY1 was induced by MeJA(methyl-jasmonic acid) and SA(salicylic acid), TwWRKY1 was induced to increase expression by MeJA in 6 hours, SA had no apparent induction in 9 hours and TwWRKY1 was induced to increase expression by SA after 12 hours. The addition of MeJA could induce the biosynthesis of triptolide, wilforgine and wilforine in T. wilfordii hairy roots using high-performance liquid chromatography (HPLC), which could apparently promote wilforgine and wilforine content and have a little effect on triptolide. The content change of triptolide, wilforgine and wilforine would be increased in early stage and decreased in late stage. The expression level of TwWRKY1 and the content change of wilforgine had the same fluctuation after addition of MeJA, it was supposed that transcription factor TwWRKY1 could regulate the secondary metabolite biosynthesis. The results will provide the scientific basis for researching the molecular characterication of TwWRKY1 gene and understanding the regulation mechanism of secondary metabolite biosynthesis pathway in T. wilfordii.
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Received: 13 October 2017
Published: 25 March 2018
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