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| Construction of a Novel Expression Vector with the Promoter of Phosphoglycerate Kinase Gene and Its Utilization of Heterogenous Gene Expression in Trichosporon fermentans |
| WANG Ping1, JIANG Mu-lan1,2, ZHANG Yin-bo1,2, WAN Xia1, LIANG Zhuo1, GONG Yang-min1 |
1. Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences, Wuhan 430062, China;
2. Key Laboratory of Biology and Genetic Improvement of Oil Crop, the Ministry of Agriculture, Wuhan 430063, China |
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Abstract Upstream sequence and partial coding sequence (CDS) of the phosphoglycerate kinase (PGK) gene were cloned from oleaginous yeast Trichosporon sp. by degenerate PCR and chromosome walking. The computation analysis revealed that a promoter-like sequence in the upstream region of the PGK coding region including essential elements such as TATA BOX and CAAT BOX was cloned. The promoter-proximal region cloned was fused with the coding region of the hygromycin gene, and then cloned into the expression vector pTF-PHT. The recombinant vector was transformed into Trichosporon fermentans, and the transformed yeast was able to grow on the selective agar containing hygromycin. The results indicated that the promoter-proximal region of the phosphoglycerate kinase gene of Trichosporon sp was able to drive the expression of heterologous genes in the Trichosporon fermentans, which provides new avenues for developing novel engineering yeast or yeast expression system.
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Received: 28 October 2011
Published: 25 March 2012
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| Fund: Supported by the National Natural Science Foundation of China(30970262, 31000778),the 863 Program(2011AA100904) and the Major S&T Projects on the Cultivation of New Varieties of Genetically Modified Organisms (Grant 2009ZX08009-120B) |
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[1] Shen J J, Li F C, Yang Q L, et al. Fermentation of Spartina anglica acid hydrolysate by Trichosporon cutaneum for microbial lipid production. Marine Sciences, 2007, 8 (31): 38-41.
[2] Wang L, Sun Y M, Qu W, et al. Effect of fermentation condition on fatty acid components of the lipid produced by Trichosporon fermentans. China Brewing, 2006, 163(10): 20-22.
[3] Fang X N, Huang W, Xia L M. Xylitol Production from corn cob hemicellulosic hydrolysate by Candida sp. Chinese Journal of Biotecnology, 2004, 2 (20): 295-298.
[4] Li X S, Lu B S, Zeng H B. Application of Trichosporon cutaneum in the production of microbial oil by enzymatic hydrolysis of distiller's grains. Liquor-Making Science & Technology, 2009, 12 (186): 87-92.
[5] Song X, Qi X Y, Qu Y B. Properties and chemical modification of lipase from Trichosporon sp. Microbiology China, 2007, 34(4): 723-726.
[6] Wu Z Q, Jia N B, Li N, et al. Construction and functional analysis of integrative expression vector in yeast. Bulletin of Biology, 2008, 43(5): 47-50.
[7] Liu X Y, Shen Y, Guo T, et al. Construction of a ribosomal DNA multi-copy integration vector and application in the industrial Saccharomyces cerevisiae strain. Journal of Shandong University (Natural Science),2005, 40 (3):105-109.
[8] Linhart C, Shamir R. The degenerate primer design problem: Theory and applications. Journal of Computational Biology, 2005, 12(4): 431-456.
[9] Hou J. Design of endonuclease restriction sites into primers for PCR cloning. Bioinformatics, 2002. 18(12): 1690-1691.
[10] Wu S X, Letchworth G J. High efficiency transformation by electroporation of Pichia pastoris pretreated with lithium acetate and dithiothreitol. BioTechniques, 2004,36(1): 152-154.
[11] Wan X, Zhang Y, Wang P, et al. Production of Gamma-Linolenic Acid in Pichia pastoris by Expression of a Delta-6 Desaturase Gene from Cunninghamella echinulata. J Microbiol Biotechnol, 2009, 19(10):1098-1102.
[12] Dyer J M, Chapital D C, Kuan J C W, et al. Production of Linolenic Acid in Yeast Cells Expressing an Omega-3 Desaturase from Tung (Aleurites fordii). JAOCS, 2004, 81(7): 647-651.
[13] Cullen D, Leong S A, Wilson L J, et al. Tansformation of Aspergillus nidulans with the hygromycin-resistance gene, hph. Gene, 1987, 57(1): 21-26.
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