| Panax saponin is the main active component of Panax ginseng(Panax ginseng C. A. Mey), which is the Chinese traditional valuable medicinal herb, it has a good medicinal value. However, the wild ginseng resource have been disrupted by excess picking, in addition, the environment for ginseng growth is harsh and saponin content is very low,so to get saponin through cultivating Panax ginseng have low effect.Now many scientists are devoting to elucidate the biosynthesis pathway of Panax saponin and study deeply on the structure and function of key enzyme gene, with the aim to improve the Panax saponin content through regulate these key enzyme gene activity by biological technology. Squalene synthase is the key enzyme in saponin biosynthesis; its quantity and activity determines the output of Panax saponin. CAS is the key enzyme of plant sterol biosynthetic pathway. CAS and DS compete common precursor2,3-oxidation squalene to make metabolization respectively flow to sterol and terpene tributarie, therefore CAS will influence the biosynthesis of saponin indirectly. RNA interference (RNAi) is phenomenon of dsRNA trigger gene silencing; introducing dsRNA into cells can induce specific mRNA degradation.In this paper, taking ginseng CAS, SQS which related to Panax saponin metabolism as research object, we amplified CAS, SQS gene interference fragment with specific primers. On the basis of principles of RNA interference technology, the interference fragment was inserted to both sides of PDK intron of pMHZ111, pMHZ112in forward and reverse way, The RNA interference vector pMHZ111-CAS, pMHZ112-CAS and pMHZ112-SQS was constructed successfully by interference fragment primer and universal primer PCR identification. Taking Agrobacterium rhizogene-mediated method to transform germfree seedling and3years old ginseng root and further induce Panax ginseng hairy root.The research results will provide foundation for further functional research of CAS, SQS in ginseng saponin biosynthetic pathway. |
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