pRS314 Plasmid


  • Model: PVT4040
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pRS314,Plasmid pRS314,pRS314 vector


pRS314 Information

Promoter: TRP1, T3, T7

Replicator: pUC ori, FI ori

Plasmid classification: yeast series, Pichia pastoris expression vector

Plasmid size: 4783bp

Prokaryotic resistance: Amp

Screening markers: TRP1

Cloned strain: DH5 alpha

Culture conditions: 37 centigrade, aerobic LB

Expression host: yeast cells

Induction method: galactose induction

5'sequencing primers: M13R:CAGGAAACAGCTATGACC

3'sequencing primers: primers designed according to sequence

Use: Yeast expression


pRS314 Description

YCtype centromere vector permitting visual detection of recombinants and production of ssDNA in E. coli. Contains promoters for in vitro RNA synthesis, priming sites useful for sequencing, and encodes the lacZ alpha (lacZ) peptide. pRSS56, constructed by ligating a PvuI fragment (bp 4982412) 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 genomic HindII/PstI fragment (1.002 kb) containing the TRP1 gene was inserted into the NdeI site and a cassette containing CEN6 and the ARS associated with histone 4 (ARSH4) was inserted into the AatII site of pRSS56. All ends were blunted. An EcoRI site in the TRP1containing fragment (external to the coding sequence) was destroyed. The order of the major features in this plasmid is: TRP1f1 ori (NaeI) T7 promoter lacZ/MCS T3 promoter pMB1 ori bla CEN6 ARSH4.




pRS314 Sequence

LOCUS       Exported                4783 bp ds-DNA     circular SYN 12-SEP-2016
DEFINITION  synthetic circular DNA
KEYWORDS    Untitled 3
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4783)
  TITLE     Direct Submission
  JOURNAL   Exported Monday, September 12, 2016 from SnapGene Viewer 3.1.4
FEATURES             Location/Qualifiers
     source          1..4783
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     misc_feature    70..573
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        610..714
                     /note="AmpR promoter"
     CDS             715..1575
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
     rep_origin      1746..2334
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
     promoter        2658..2688
                     /note="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    2696..2712
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="lac operator"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     2720..2736
                     /note="M13 rev"
                     /note="common sequencing primer, one of multiple similar 
     CDS             2732..3307
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
     promoter        2757..2775
                     /note="T3 promoter"
                     /note="promoter for bacteriophage T3 RNA polymerase"
     misc_feature    2788..2895
                     /note="pBluescript multiple cloning site"
     primer_bind     2805..2821
                     /note="KS primer"
                     /note="common sequencing primer, one of multiple similar 
     primer_bind     complement(2855..2871)
                     /note="SK primer"
                     /note="common sequencing primer, one of multiple similar 
     promoter        complement(2904..2922)
                     /note="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2929..2945)
                     /note="M13 fwd"
                     /note="common sequencing primer, one of multiple similar 
     rep_origin      3090..3545
                     /note="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     CDS             complement(3642..4316)
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /note="yeast auxotrophic marker"
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