Over - expression effect of gene encoding 3-hydroxy-3-methylglutaryl-CoA reductase on production of taxol in Iranian hazel [Corylus avellana L.]

ABSTRACT

Sustainable and commercial production of taxol as an anti cancer drug is a critical point to its clinical application. Nowadays, hazel because of rapid growth and wide range distribution is considered as an alternative source of Taxol. To increase taxol production the cDNA encoding 3-hydroxy-3-methylglutaryl-CoA reductase [HMGR] from Iranian hazel [GeneBank accession number KF306244, showed by CIHMGR] was isolated and over-expressed in pCAMBIA1304 binary vector. The effect of transient over-expression of HMGR in callus and leaf were evaluated on Taxol production. The calli was established through the culture of immature cotyledon on Murashige and Skoog basal medium supplemented with 2, 4-D and BA. The first strand cDNA of CiHMGR was synthesized by specific primers. Enzymatic assay of recombinant CiHMGR in E. coli were done by western blott and His-tag affinity techniques. Also production of taxol in transformed callus and leaf were evaluated by HPLC analysis. An Open Reading Frame [ORF] with 1698 bp length and a deduced polypeptide with 566 amino acid residues were amplified. The highest and lowest amount of taxol was 0.016 mg/g. DW and 0.004 mg/gDW in transformed calli and untransformed leaves respectively. Generally the over-expression of HMGR increase the total isoprenoids yield, therefore to have high production of target secondary metabolites [taxol] we need both of network of transformed genes and elicited cell culture