Size (kb): 8.81
Vector: pMMB66EH (plasmid)
Promoters: Promoter tac
Construction: RSF1010, pKK223-3, M13mp8 polylinker, lacIq
Replicon: RSF1010, oriV, oriT
Repressor gene: lacIq
Transcription terminator: rrnB
Contains the following sites separated by (bp): EcoRI-764 -AhaIII-97 -ScaI-112 -PvuI-2101 -AccI-1848 -ScaI-55 -AccI-2039 -BstEII-372 -PvuII-93 -PvuII-94 -HpaI-320 -BstEII-182 -MluI-736 -EcoRI.
Restriction digests of the clone give the following sizes (kb): AccI--3.9, 3.0, 1.9; BamHI--8.8; HindIII/PvuII--7.6, 1.3.
In the pMMB66 (ATCC 37620
, 37621) and pMMB67 (ATCC 37622
, 37623) vector series, the EH and HE in the name designate the orientation of the multiple cloning site to the tac promoter.
An autoregulated high-level expression vector utilizing the tac promoter, rrnB terminators and lacIq for lac repression. Concentration of induced gene product may be regulated by varying the lac inducer concentration.
Can be mobilized by conjugative IncP helper plasmids [e.g. pRK2013 (ATCC 37159)
, pUB307] into Klebsiella aerogenes, Proteus mirabilis, Pseudomonas aeruginosa, Serratia marcescens, and other gram-negative bacteria.
The sequence of RSF1010 is given in this reference.
A 1246 bp lacIQ HindIII fragment was inserted at XmaIII/BamHI of pKK223-3. The 2995 bp NruI/AhaIII fragment of the resulting plasmid was cloned between PstI (bp 7768) and PvuII (bp 1951) of RSF1010.
Furste JP, et al. Molecular cloning of the plasmid RP4 primase region in a multi-host-range tacP expression vector. Gene 48: 119-131, 1986. PubMed: 3549457
Scholz P, et al. Complete nucleotide sequence and gene organization of the broad-host-range plasmid RSF1010. Gene 75: 271-288, 1989. PubMed: 2653965
Erich Lanka, personal communication
Frozen glycerol stock of E. coli containing the plasmid