pMLP9 (ATCC® 87445)

Applications: vector permitting positive selection for inserts  /  Depositors: F Puta

Permits and Restrictions

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Designations pMLP9
Depositors F Puta
Biosafety Level 1
Vector Information
Size (kb): 4.7899999618530270
Vector: pMLP9 (plasmid)
Promoters: Promoter lambda PR
Construction: pMLP21, pML25
Marker(s):ampR,cmlR
Construct size (kb): 4.789999961853027
Features: marker(s): ampR
marker(s): cmlR
promoter: lambda PR
promoter: tetR
replicon: pMB1
repressor gene: cI
transcription terminator: tandem fd terminators
Applications
vector permitting positive selection for inserts
Comments
Restriction digests of the clone give the following sizes (kb): HindIII--4.8; EcoRI--4.0, 0.9; BamHI--4.8.
pMLP9 is the same as pMLP8, except that a BglII site has been added to pMLP9.
The cytoplasmic chloramphenicol resistance protein does not negatively affect cell growth when overproduced (in contrast to cloning vectors that overexpress membrane associated tetR protein).
The level of resistance to chloramphenicol differs substantially depending on the stage at which they are tested.
The chloramphenicol resistance of the plasmid should be tested before cloning inserts.
This is a postive selection cloning vector which will confer chloramphenicol resistance (recommended concentration is 10ug/ml) only when an insert has been successfully cloned into the cI repressor gene.
Recommendation for verfication: HindIII; BglII; SmaI.
A pair of phage fd rho-independent strong terminators downstream of the cmlR gene prevents transcription through the origin of replication. This stabilizes recombinant plasmid copy number when the PR promoter for the cmlR gene is derepressed.
Media Medium 1227: LB Medium (ATCC medium 1065) with 50 mcg/ml ampicillin
Growth Conditions
Temperature: 37.0°C
References

Puta F, et al. Direct selection shuttle plasmid vector, pPW264, used for cloning the alpha-amylase gene of Schwanniomyces occidentalis. Folia Microbiol. 39: 255-260, 1994. PubMed: 7729761

Frantisek Puta, personal communication

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