Construction of a Rhizopus arrhizus glucoamylase gene suitable for expression in distinct host: introns spliced artificially by PCR.

Construction of a Rhizopus arrhizus glucoamylase gene suitable for expression in distinct host: introns spliced artificially by PCR. Research Abstract Details 

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Construction of a Rhizopus arrhizus glucoamylase gene suitable for expression in distinct host: introns spliced artificially by PCR. Abstract Text:

Liquan Yang,Xiaojun Dai,Jianhua Hou,Chunxiao Ma,Cuiyan Wang,Zhiqiang Wu,Minggang Li,Liquan Yang,Xiaojun Dai,Jianhua Hou,Chunxiao Ma,Cuiyan Wang,Zhiqiang Wu,Minggang Li,

Glucoamylase is an industrially extremely important enzyme in the fermentative production of ethanol, used in the enzymatic conversion of starch into high glucose and fructose syrups. The aim of this study is to construct a Rhizopus arrhizus glucoamylase gene (RaGA)-introns artificially spliced by PCR-suitable for expression in S. cerevisiae host and tried expressing in Picha pastoris. In previous work, we failed in amplifying glucoamylase gene from R. arrhizus by RT-PCR, so several primers were designed to splicing the introns by PCR in vitro. Sequence analysis shown that all introns in the RaGA were deleted correctly and no mutant was induced in the extrons compared with the RaGA gene originally cloned. The RaGA gene artificially constructed was transferred into P. pastoris integrative expression vectors pPIC9 (containing small a, Cyrillic-factor). Consequently, the plasmids pPIC9-RaGA was lineared by SacI and inserted into P. pastoris GS115 (His(-)) genome downstream of the 5'AOX1 promoter by the method of electroporation. Induction by 0.75% methanol for 72 h led to synthesis of secreted glucoamylase. So it is demonstrated that the glucoamylase gene has been expressed in and secreted from P. pastoris.

Construction of a Rhizopus arrhizus glucoamylase gene suitable for expression in distinct host: introns spliced artificially by PCR. Publishing Authors By Initials

L Yang,X Dai,J Hou,C Ma,C Wang,Z Wu,M Li,L Yang,X Dai,J Hou,C Ma,C Wang,Z Wu,M Li,

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Construction of a Rhizopus arrhizus glucoamylase gene suitable for expression in distinct host: introns spliced artificially by PCR. Journal Published:

PUBLICATION TYPE: Journal Article

Journal: Molecular biology reports

VOLUME: 35

Page Numbers: 9-15

Journal Abbreviation: Mol. Biol. Rep.

ISSN: 0301-4851

DAY: 20

MONTH: 06

YEAR: 2007

Construction of a Rhizopus arrhizus glucoamylase gene suitable for expression in distinct host: introns spliced artificially by PCR. Information

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LANGUAGE: eng

NlmUniqueID: 403234

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Grant and Affiliation Information for Construction of a Rhizopus arrhizus glucoamylase gene suitable for expression in distinct host: introns spliced artificially by PCR.

AFFILIATION: Tianjin Key Laboratory of Microbial Functional Genomics, The Key Laboratory of Bioactive Material, Ministry of Education, Tianjin, 300071, China.

Country: Netherlands

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MEDLINETA: Mol Biol Rep

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