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Construction of an Arxula adeninivorans host-vector system based on trp1 complementation

Paper ID Volume ID Publish Year Pages File Format Full-Text
25478 43576 2007 10 PDF Available
Title
Construction of an Arxula adeninivorans host-vector system based on trp1 complementation
Abstract

A host/vector expression system based on an Arxula adeninivorans Δatrp1 gene disruption mutant has been constructed. For this purpose the ATRP1 gene encoding a phosphoribosyl anthranilate isomerase was isolated from the yeast A. adeninivorans and its genome locus was characterized. The Δatrp1 mutant was generated applying an amplified DNA fragment containing the ALEU2m gene flanked by ATRP1 gene sequences of some 750 bp. The generated auxotrophic host strain was transformed with the plasmid pAL-ATRP1-amyA, which contains the ATRP1 gene as selection marker and the 25S rDNA for targeting. For expression assessment, the plasmid was equipped with an expression cassette consisting of the Bacillus amyloliquefaciens-derived amyA gene fused to the constitutive A. adeninivorans-derived TEF1 promoter and Saccharomyces cerevisiae-derived PHO5 terminator. Transformants contained a single chromosomal copy of the heterologous DNA and were found to be mitotically stable. In initial fermentation trials on a 200 ml shake flask scale maximal α-amylase product levels of ca. 300 nkat ml−1 were observed after 72 h of cultivation with more than 95% of the recombinant α-amylase accumulated in the culture medium.

Keywords
Arxula adeninivorans; Auxotrophic selection marker; ATRP1 gene; Gene disruption; Heterologous gene expression; Transformation
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Construction of an Arxula adeninivorans host-vector system based on trp1 complementation
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 127, Issue 3, 10 January 2007, Pages 392–401
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us