Abstract:
:The Red recombinase system, the most convenient genetic tool applied in Escherichia coli and other bacteria, was introduced for gene replacement in Klebsiella pneumoniae. The novel K. pneumoniae gene replacement system comprised the Red and FLP recombinases expression vector pDK6-red and pDK6-flp, and linear DNA fragments which encompassed a selective marker gene with target gene flanking extensions; the latter were PCR amplified using a plasmid DNA template obtained by in vivo recombination in E. coli. In this study, dhak1 gene, encoding a subunit of dihydroxyacetone kinase II, was deleted markerlessly at a transformation ratio of 260 CFU/μg DNA, i.e., 1,000-fold higher than that achieved in the native way. Our studies provide an efficient method with detailed protocol to perform gene replacement in K. pneumoniae and has great potential to be developed as a routine genetic approach for this important industrial microorganism.
journal_name
J Ind Microbiol Biotechnoljournal_title
Journal of industrial microbiology & biotechnologyauthors
Wei D,Wang M,Shi J,Hao Jdoi
10.1007/s10295-012-1117-xsubject
Has Abstractpub_date
2012-08-01 00:00:00pages
1219-26issue
8eissn
1367-5435issn
1476-5535journal_volume
39pub_type
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