pSLF373 (ATCC® 87614)

Applications: encodes an epitope tag for protein isolation or monitoringexpression vector  /  Depositors: SL Forsburg

Permits and Restrictions

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Designations pSLF373
Depositors SL Forsburg
Biosafety Level 1
Vector Information
Size (kb): 8.5000000000000000
Vector: pSLF373 (phagemid)
Promoters: Promoter for expression nmt1** (low strength; nmt1)
Construction: REP82X
Marker(s):ampR,ura4+
Construct size (kb): 8.5
Features: marker(s): ampR
marker(s): ura4+
promoter for expression: nmt1** (low strength; nmt1)
replicon: ars1
replicon: pMB1, f1
MCS: XhoI...SmaI
epitope tag (N-terminal): hemagglutinin (HA) triple tag
Applications
encodes an epitope tag for protein isolation or monitoring
expression vector
Comments
Restriction digests of the clone give the following sizes (kb): HindIII--5.6, 1.7, 1.0; EcoRI--7.4, 1.2; BamHI--8.5.
The fission yeast tagging vectors, pSLF173 (ATCC 87612), pSLF273 (ATCC 87613) and pSLF373 (ATCC 87614), contain three versions of the nmt1 promoter: full strength (nmt1), medium strength (nmt1*) and low strength (nmt1**), respectively.
The weaker promoters (nmt1* and nmt1**) contain mutations that attenuate both repressed and induced levels of expression.
Each version of the nmt1 promoter can be expressed at low or high levels in thiamine-free media.
The vector was designed to tag expressed protein at N-terminus with triple HA tag, which contains an internal BamHI site. The vector lack a stop codon.
The vector was constructed by cutting pSLF173 with SacI and XhoI, isolating the fragment containing the HA polylinker, and ligating into XhoI-SacI digested REP82X.
Media ATCC® Medium 1227: LB Medium (ATCC medium 1065) with 50 mcg/ml ampicillin
Growth Conditions
Temperature: 37.0°C
References

Forsburg SL, Sherman DA. General purpose tagging vectors for fission yeast. Gene 191: 191-195, 1997. PubMed: 9218719

Basi G, et al. TATA box mutations in the Schizosaccharomyces pombe nmt1 promoter affect transcription efficiency but not the transcription start point or thiamine repressibility. Gene 123: 131-131, 1993. PubMed: 8422997

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