pRS166 (ATCC® 77149)

Applications: YC-type (centromeric) shuttle vectorYX-type (expression) shuttle vectorexpression vectorshuttle vectorvector containing primer sites useful for sequencingvector permitting RNA synthesis in vitrovector permitting construction of fusion proteinsvector permitting production of single-stranded DNAvector permitting visual detection of recombinants  /  Depositors: P Hieter

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Designations pRS166
Depositors P Hieter
Biosafety Level 1
Vector Information
Size (kb): 6.3000001907348630
Vector: pRS166 (phagemid)
Promoters: Promoter T3
Construction: pRSS56 [pBluescript KS+, pBS(+)], GAL1/10 promoter
Marker(s):TRP1,ampR
Construct size (kb): 6.300000190734863
Features: insert detection: lacZ'
marker(s): ampR, TRP1
promoter: lac, T3, T7, GAL10, URA3
replicon: pMB1, f1, ARS1
Applications
YC-type (centromeric) shuttle vector
YX-type (expression) shuttle vector
expression vector
shuttle vector
vector containing primer sites useful for sequencing
vector permitting RNA synthesis in vitro
vector permitting construction of fusion proteins
vector permitting production of single-stranded DNA
vector permitting visual detection of recombinants
Comments
Restriction digests of the clone give the following sizes (kb): PvuI--3.6, 3.0; SalI--6.6; EcoRI--6.6; SmaI--6.6; BamHI--6.6.
Expression is inducible by galactose.
YX-type expression shuttle vector permitting visual detection of recombinants and production of ssDNA in E. coli. Contains promoters for in vitro RNA synthesis and encodes the lacZ alpha (lacZ') peptide.
Transcription from the GAL10 promoter (ATG is included in the sequence) permits the synthesis of a fusion protein (starting with 26 aa of GAL10) through the polylinker in the same frame as lacZ'.
pRSS56, constructed by ligating a PvuI fragment (bp 498-2412) of pBluescript KS+ to a PvuI fragment (bp 2850-730) of pBS(+), contains the KS MCS from pBluescript KS+ and the unique NdeI and AatII sites between bla and f1 origin of pBS(+).
A fragment containing the TRP1 gene was inserted into the NdeI site, a DraIII/EcoRI fragment containing the GAL10 promoter was inserted into the Asp718 site and a URA3 fragment was inserted between the SpeI and NotI sites of the MCS.
The order of the major features in this plasmid is: TRP1 - URA 3 - f1 ori (NaeI) - T7 promoter - lacZ'/MCS - T3 promoter - GAL10 promoter - GAL1 promoter - pMB1 ori - bla - CEN - ARS1.
Media ATCC® Medium 1227: LB Medium (ATCC medium 1065) with 50 mcg/ml ampicillin
Growth Conditions
Temperature: 37.0°C
References

Philip Hieter, personal communication

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