Vector: pNJ17

Number of plasmids with this backbone: 1
Name: pNJ17
Description: Vector for transposon-based mutagenesis in bacteria, with arabinose-inducible promoter to drive expression of gene adjacent to transposon insert (modified promoter-out approach); kanamycin resistance; restriction enzyme cloning.
Synonyms: TnAraOut
Type: bacterial plasmid
Form: dsDNA
Size (bp):: 6722
Properties: inducible expression, mutagenesis vector, transposon mediated gene disruption
Vector Map: pNJ17.pdf
Vector Sequence: pNJ17.txt
Comments: This shipment includes the following items: 1) Purified pNJ17 (Judson and Mekalanos, 2000. PMID: 10888841). 2) E. coli strain DH5alpha lambda pir. 3) E. coli strain BW20767 (Metcalf, et al., 1996. PMID: 8693022). It is recommended that the following approaches be used for handling this plasmid: 1) Avoid single colony isolation for strains containing pNJ17. Instead, it is preferable to scrape up multiple individual colonies from a plate for establishing stocks and cultures. Strain passage should be minimized. This is because the transposon hops out at a relatively high frequency, causing plasmid loss. Plasmid loss cannot be tracked by kanamycin resistance because insertion of the transposon into the host strain chromosome should also cause a resistant phenotype. Please also keep in mind that mariner-containing plasmids can be prone to spontaneous rearrangement. 2) DH5alpha lambda pir is suitable for cloning or plasmid maintenance, but it is not competent to mobilize the plasmid into recipient strains. For mating, the plasmid should be introduced into BW20767 or another mating strain of your choice. Multi-colony isolation should also be used for the mating strains once the plasmid has been introduced.

Features:

Type Name Description Start Position End Position
transposon fragment transposase mariner-type transposase 0 0
transposon fragment inverted repeat (1) transposon inverted repeat 0 0
transposon fragment inverted repeat (2) transposon inverted repeat 0 0
promoter pBAD pr pBAD promoter (arabinose-inducible) oriented outward (toward Tn inverted repeat prior to insertion) for induction of adjacent gene after insertion 0 0
MCS MCS MCS (SalI, XbaI, KpnI, EcoRI, NheI) 0 0
gene fragment RP4 RP4 (oriT) 0 0
selectable marker kanR kanamycin resistance gene 0 0
bacterial origin oriR6K-gamma oriR6K-gamma for bacterial conjugation 0 0
gene fragment araC araC gene 0 0

Authors:

Author Name Author Type Creation Date
John Mekalanos Academic Researcher, vector PI
Su Chiang Academic Researcher, vector donor
Nicholas Judson Academic Researcher, vector first author

Publications:

PMID Title
10888841 TnAraOut, a transposon-based approach to identify and characterize essential bacterial genes

Parent Vector:

Name Comments