PGP1331

pGP1331: click to enlarge

General comments

Integrative plasmid for the fusion of 3x FLAG tag to the C-terminus of a protein, keeping the control of the expression under its natural promotor

• E.coli: multicopy plasmid, select for ampicillin
• B. subtilis: no ori, select for spectinomycin

Hints for the usage

• amplify the last 600bp of your gene of interest,
• skip the Stop-codon of the gene in the reverse primer
• take care that the integration is in-frame
• Don't use XbaI and KpnI for cloning. They cut the vector twice!

Antibody

• dilute the anitbody 1:10000 (you can freeze the Blotto solution and reuse it)

The reference

Martin Lehnik-Habrink, Henrike Pförtner, Leonie Rempeters, Nico Pietack, Christina Herzberg, Jörg Stülke
The RNA degradosome in Bacillus subtilis: identification of CshA as the major RNA helicase in the multiprotein complex.
Mol Microbiol: 2010, 77(4);958-71
[PubMed:20572937] [WorldCat.org] [DOI] (I p)