Biology Lab Report

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Study area: Present study is based on laboratory work; however the plant material for experimental purpose will be collected from experimental farm of CSIR-IIIM (32.73N and 74.87E). Objectives: 1. Full length cloning of squalene synthase, squalene epoxidase, β- amyrin synthase CYP88D6, CYP172A54, the five known genes involved in glycyrrhizin biosynthesis from Glycyrrhiza glabra. 2. Real time expression analyses of the targeted genes in various in-vitro regenerated plant organs in relation to organ specific glycyrrhizin accumulation. 3. Phytochemical analysis of in vitro regenerated plants. 4. Functional and biochemical characterization of at least one protein expressed in heterologous plant host. REVIEW OF LITERATURE The…show more content…
Biosynthesis of glycyrrhizin is not fully elucidated at gene level. The early stages of triterpenoid saponin biosynthesis via isoprenoid pathway involve the dimerization of two molecules of farnesyl diphosphate catalyzed by squalene synthase, epoxidase–mediated oxidation then produces 2,3-oxidosqualene. The first diversifying step in triterpenoid biosynthesis is the cyclization of 2,3-oxidosqualene a common substrate of oxidosqualene cyclases (OSCs) and a precursor of both triterpenes and sterols [Abe et al., 1993]. The biosynthesis of glycyrrhizin involves the initial cyclization of 2, 3-oxidosqualene by a specific OSC, β-amyrin synthase (bAS), to the primarily oleanane triterpene β-amyrin which is one of the most commonly occurring triterpenes in plants. The subsequent steps involve a series of oxidative reactions at positions C-11 (two-step oxidation) and C-30 (three-step oxidation), followed by glycosyl transfers to the C-3 hydroxyl group. Genes involved in the early stages of glycyrrhizin biosynthesis, namely, squalene synthase and bAS, have been functionally isolated from G. glabra [Hayashi et al., 1999, 2001]. The bAS gene has also been functionally isolated from several other plants including Arabidopsis thaliana [Shibuya et al., 2009], Avena strigosa [Qi et al., 2004], Lotus japonicus [Sawai et al., 2006], and Medicago truncatula [Suzuki et al., 2002]; however, most of the steps in the modification of the

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