New selectable host-marker systems for multiple genetic manipulations based on TRP1, MET2, and ADE2 in the methylotrophic yeast Hansenula polymorpha = 메탄올자화효모 한세눌라 폴리모파의 다중유전자 조작을 위한 신규 선별 숙주-표지 유전자 시스템

Cited 16 time in scopus
Metadata Downloads
Title
New selectable host-marker systems for multiple genetic manipulations based on TRP1, MET2, and ADE2 in the methylotrophic yeast Hansenula polymorpha = 메탄올자화효모 한세눌라 폴리모파의 다중유전자 조작을 위한 신규 선별 숙주-표지 유전자 시스템
Author(s)
S A Cheon; J Choo; V M Ubiyvovk; J N Park; Moo Woong Kim; Doo-Byoung OhOh Suk Kwon; A A Sibirny; J Y Kim; H A Kang
Bibliographic Citation
Yeast, vol. 26, no. 9, pp. 507-521
Publication Year
2009
Abstract
Interest has been increasing in the thermotolerant methylotrophic yeast Hansenula polymorpha as a useful system for fundamental research and applied purposes. Only a few genetic marker genes and auxotrophic hosts are yet available for this yeast. Here we isolated and developed H. polymorpha TRP1, MET2 and ADE2 genes as selectable markers for multiple genetic manipulations. The H. polymorpha TRP1 (HpTRP1), MET2 (HpMET2) and ADE2 (HpADE2) genes were sequentially disrupted, using an HpURA3 pop-out cassette in H. polymorpha to generate a series of new multiple auxotrophic strains, including up to a quintuple auxotrophic strain. Unexpectedly, the HpTRP1 deletion mutants required additional tryptophan supplementation for their full growth, even on complex media such as YPD. Despite the clearly increased resistance to 5-fluoroanthranilic acid of the HpTRP1 deletion mutants, the HpTRP1 blaster cassette does not appear to be usable as a counter-selection marker in H. polymorpha. Expression vectors carrying HpADE2, HpTRP1 or HpMET2 with their own promoters and terminators as selectable markers were constructed and used to co-transform the quintuple auxotrophic strain for the targeted expression of a heterologous gene, Aspergillus saitoi MsdS, at the ER, the Golgi and the cell surface, respectively. The nucleotide sequences presented here were submitted to GenBank under Accession Nos AY795576 (HpTRP1), FJ226453 (HpMET2) and FJ493241 (HpADE2), respectively.
Keyword
ADE2Auxotrophic markerExpression vectorHansenula polymorphaMET2TRP1
ISSN
0749-503X
Publisher
Wiley
DOI
http://dx.doi.org/10.1002/yea.1701
Type
Article
Appears in Collections:
Division of Bio Technology Innovation > SME Support Center > 1. Journal Articles
Files in This Item:
  • There are no files associated with this item.


Items in OpenAccess@KRIBB are protected by copyright, with all rights reserved, unless otherwise indicated.