pSLF173 (ATCC® 87612)

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

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Designations pSLF173
Depositors SL Forsburg
Biosafety Level 1

Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country.

Vector Information
Size (kb): 8.5000000000000000
Vector: pSLF173 (phagemid)
Promoters: Promoter for expression nmt1 (full strength)
Construction: REP4X
Construct size (kb): 8.5
Features: marker(s): ampR
marker(s): ura4+
promoter for expression: nmt1 (full strength)
replicon: ars1
replicon: f1
replicon: pMB1
MCS: XhoI...SmaI
epitope tag (N-terminal): hemagglutinin (HA) triple tag
encodes an epitope tag for protein isolation or monitoring
expression vector
Restriction digests of the clone give the following sizes (kb): BamHI--8.5; EcoRI--7.3, 1.2; PstI--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 1) amplification by PCR with primers designed to flank the triple HA tag in Bluescript-HA and to modify the polylinker, 2) gel purification of the PCR product and digest with XhoI
and 3) ligation into REP4X cleaved with XhoI and SmaI.
Media ATCC® Medium 1227: LB Medium (ATCC medium 1065) with 50 mcg/ml ampicillin
Growth Conditions
Temperature: 37.0°C

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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