Transcription Factors Analysis Involved In Saikosaponins Biosynthesis Of Bupleurum Chinense DC. Based On The Promoter Interactants And Synchronicity Of BcAS1

Bupleurum chinense DC., is known as one major source of Radix Bupleuri, with triterpenoid saponins-saikosaponins as its major bioactive components. In triterpene biosynthetic pathway,2,3-oxidosqualene leads to production of oleanane-type saikosaponins through the activity of β-AS. However, few TF genes which regulate β-AS expression have been documented. Therefore, this study cloned the promoter of BcAS1 and selected the related TF gene by yeast one-hybrid experiments; moreover, we screened some candidate TF genes by comparing with β-AS expression pattern. Our results lay a valuable foundation for saikosaponins biosynthesis transcriptional regulation.Specific findings are as follows:1. A 783 bp fragment upstream the start codon of BcASl in B. chinense was obtained. According to BcAS1 cDNA, the full-length gene of BcASl was acquired. BcASl is 6019 bp long which encodes 764 amino acid residues and contains 17 introns. The transient expression in the onion epidermal cells showed that the promoter sequence of BcASl gene was conducted successfully. The the functional elements of BcASl promoter were analyzed by the bioinformatics analysis. The result showed that the sequence contained the some dry-resistant elements (MBS、HSE)、 some important cis-regulatory elements, such as ABRE、TCA-element, and some relation to light elements are also found.2. The results of the analysis of cis-acting elements, select ABRE element at the same direction as the tandern repeat 3 times bait vector construct the target sequence. Using yeast one-hybrid technology, the B. chinense adventitious roots cDNA library was screened and one TF-BcTF5 was detected belonging to bZIP family.3. A total of 27 AP2 family TFs were selected for MeJA inducibility and tissue-specific expression patterns analysis using real-time PCR. And we analyzed the relationship to the saikosaponins accumulation in MeJA-treated adventitious roots and β-AS tissue specific expression patterns. Two AP2 family TFs were identified as the most likely candidates involved in saikosaponin biosynthesis.