NDLI: Development of a New Generation of Vectors for Gene Expression, Gene Replacement, and Protein-Protein Interaction Studies in Mycobacteria

Content Provider	PubMed Central
Author	Parikh, Amit Kumar, Devanand Chawla, Yogesh Krishna, Kurthkoti Khan, Shazia Varshney, Umesh Nandicoori, Vinay K.
Copyright Year	2013
Abstract	Escherichia coli-mycobacterium shuttle vectors are important tools for gene expression and gene replacement in mycobacteria. However, most of the currently available vectors are limited in their use because of the lack of extended multiple cloning sites (MCSs) and convenience of appending an epitope tag(s) to the cloned open reading frames (ORFs). Here we report a new series of vectors that allow for the constitutive and regulatable expression of proteins, appended with peptide tag sequences at their N and C termini, respectively. The applicability of these vectors is demonstrated by the constitutive and induced expression of the Mycobacterium tuberculosis pknK gene, coding for protein kinase K, a serine-threonine protein kinase. Furthermore, a suicide plasmid with expanded MCS for creating gene replacements, a plasmid for chromosomal integrations at the commonly used L5 attB site, and a hypoxia-responsive vector, for expression of a gene(s) under hypoxic conditions that mimic latency, have also been created. Additionally, we have created a vector for the coexpression of two proteins controlled by two independent promoters, with each protein being in fusion with a different tag. The shuttle vectors developed in the present study are excellent tools for the analysis of gene function in mycobacteria and are a valuable addition to the existing repertoire of vectors for mycobacterial research.
Related Links	http://dx.doi.org/10.1128/aem.03695-12
Ending Page	1729
Page Count	12
Starting Page	1718
File Format	PDF
ISSN	00992240
e-ISSN	10985336
Journal	Applied and Environmental Microbiology
Issue Number	5
Volume Number	79
Language	English
Publisher	American Society for Microbiology
Publisher Date	2013-03-01
Access Restriction	Open
Rights Holder	American Society for Microbiology
Subject Keyword	Biotechnology Food Science Ecology Applied Microbiology and Biotechnology Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Ecology Food Science Applied Microbiology and Biotechnology Biotechnology

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