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Sequential gene integration for the engineering of Kluyveromyces marxianus

Paper ID Volume ID Publish Year Pages File Format Full-Text
25480 43576 2007 9 PDF Available
Title
Sequential gene integration for the engineering of Kluyveromyces marxianus
Abstract

The attributes of the yeast Kluyveromyces marxianus (rapid growth rate at high temperature, utilization of a wide range of inexpensive carbon sources) make it a promising industrial host for the synthesis of protein and non-protein products. However, no stable multicopy plasmids are currently available for long-term culture of K. marxianus. To allow the stable genetic/metabolic engineering of K. marxianus, a method for integrating precise numbers of the same or different genes was developed for this yeast. A K. marxianus URA3 deletion mutant was constructed and the URA3 blaster (UB) reusable selection cassette from Saccharomyces cerevisiae was used to select sequential, untargeted chromosomal insertions of the Bacillus megaterium lactate dehydrogenase (LDH) gene. Following excision of the UB cassette from the chromosomes, the integrating vector was retransformed into the strain and a second copy of LDH was inserted, demonstrating the success of this method for sequential gene integrations in K. marxianus. LDH activity and lactic acid concentration increased with each gene insertion, further illustrating the success of this method.

Keywords
Kluyveromyces marxianus; Gene integration; Lactic acid; Lactate dehydrogenase
First Page Preview
Sequential gene integration for the engineering of Kluyveromyces marxianus
Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 127, Issue 3, 10 January 2007, Pages 408–416
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering