pRS314

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  • Model: PVT4040
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pRS314

PVT4040   2ug
 

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-vector

 

 

pRS314 Sequence

LOCUS       Exported                4783 bp ds-DNA     circular SYN 12-SEP-2016

DEFINITION  synthetic circular DNA

ACCESSION   .

VERSION     .

KEYWORDS    Untitled 3

SOURCE      synthetic DNA construct

  ORGANISM  synthetic DNA construct

REFERENCE   1  (bases 1 to 4783)

  AUTHORS   .

  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="CEN/ARS"

                     /note="S. cerevisiae CEN6 centromere fused to an 

                     autonomously replicating sequence"

     promoter        610..714

                     /gene="bla"

                     /note="AmpR promoter"

     CDS             715..1575

                     /codon_start=1

                     /gene="bla"

                     /product="beta-lactamase"

                     /note="AmpR"

                     /note="confers resistance to ampicillin, carbenicillin, and

                     related antibiotics"

                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI

                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS

                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFFHNMGDHVTRLDRW

                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA

                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS

                     LIKHW"

     rep_origin      1746..2334

                     /direction=RIGHT

                     /note="ori"

                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 

                     replication"

     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 

                     variants"

     CDS             2732..3307

                     /codon_start=1

                     /gene="lacZ fragment"

                     /product="LacZ-alpha fragment of beta-galactosidase"

                     /note="lacZ-alpha"

                     /translation="MTMITPSSELTLTKGNKSWVPGPPSRSTVSISLISNSCSPGDPLV

                     LERPPPRWSSNSPYSESYYNSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART

                     DRPSQQLRSLNGEWRDAPCSGALSAAGVVVTRSVTATLASALAPAPFAFFPSFLATFAG

                     FPRQALNRGLPLGFRFSALRHLDPKKLD"

     promoter        2757..2775

                     /note="T3 promoter"

                     /note="promoter for bacteriophage T3 RNA polymerase"

     misc_feature    2788..2895

                     /note="MCS"

                     /note="pBluescript multiple cloning site"

     primer_bind     2805..2821

                     /note="KS primer"

                     /note="common sequencing primer, one of multiple similar 

                     variants"

     primer_bind     complement(2855..2871)

                     /note="SK primer"

                     /note="common sequencing primer, one of multiple similar 

                     variants"

     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 

                     variants"

     rep_origin      3090..3545

                     /direction=RIGHT

                     /note="f1 ori"

                     /note="f1 bacteriophage origin of replication; arrow 

                     indicates direction of (+) strand synthesis"

     CDS             complement(3642..4316)

                     /codon_start=1

                     /gene="S. cerevisiae TRP1"

                     /product="phosphoribosylanthranilate isomerase, required 

                     for tryptophan biosynthesis"

                     /note="TRP1"

                     /note="yeast auxotrophic marker"

                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK

                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES

                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW

                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA

                     KK"

     promoter        complement(4317..4597)

                     /gene="S. cerevisiae TRP1"

                     /note="TRP1 promoter"

ORIGIN

        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt

       61 cttaggacgg atcgcttgcc tgtaacttac acgcgcctcg tatcttttaa tgatggaata

      121 atttgggaat ttactctgtg tttatttatt tttatgtttt gtatttggat tttagaaagt

      181 aaataaagaa ggtagaagag ttacggaatg aagaaaaaaa aataaacaaa ggtttaaaaa

      241 atttcaacaa aaagcgtact ttacatatat atttattaga caagaaaagc agattaaata

      301 gatatacatt cgattaacga taagtaaaat gtaaaatcac aggattttcg tgtgtggtct

      361 tctacacaga caagatgaaa caattcggca ttaatacctg agagcaggaa gagcaagata

      421 aaaggtagta tttgttggcg atccccctag agtcttttac atcttcggaa aacaaaaact

      481 attttttctt taatttcttt ttttactttc tatttttaat ttatatattt atattaaaaa

      541 atttaaatta taattatttt tatagcacgt gatgaaaagg acccaggtgg cacttttcgg

      601 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg

      661 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt

      721 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt

      781 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg

      841 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa

      901 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt

      961 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag

     1021 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt

     1081 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga

     1141 ccgaaggagc taaccgcttt ttttcacaac atgggggatc atgtaactcg ccttgatcgt

     1201 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta

     1261 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg

     1321 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc

     1381 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt

     1441 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg

     1501 ggcagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg

     1561 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa

     1621 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa

     1681 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga

     1741 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg

     1801 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact

     1861 ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac

     1921 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg

     1981 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg

     2041 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga

     2101 acgacctaca ccgaactgag atacctacag cgtgagcatt gagaaagcgc cacgcttccc

     2161 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg

     2221 agggagcttc caggggggaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc

     2281 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc cgagcctatg gaaaaacgcc

     2341 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt

     2401 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc

     2461 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc

     2521 ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac

     2581 aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag ttacctcact

     2641 cattaggcac cccaggcttt acactttatg cttccggctc ctatgttgtg tggaattgtg

     2701 agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa gctcggaatt

     2761 aaccctcact aaagggaaca aaagctgggt accgggcccc ccctcgaggt cgacggtatc

     2821 gataagcttg atatcgaatt cctgcagccc gggggatcca ctagttctag agcggccgcc

     2881 accgcggtgg agctccaatt cgccctatag tgagtcgtat tacaattcac tggccgtcgt

     2941 tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca

     3001 tccccccttc gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca

     3061 gttgcgcagc ctgaatggcg aatggcgcga cgcgccctgt agcggcgcat taagcgcggc

     3121 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc

     3181 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa

     3241 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact

     3301 tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt

     3361 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa

     3421 ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt

     3481 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgtttac

     3541 aatttcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcaggcaa

     3601 gtgcacaaac aatacttaaa taaatactac tcagtaataa cctatttctt agcatttttg

     3661 acgaaatttg ctattttgtt agagtctttt acaccatttg tctccacacc tccgcttaca

     3721 tcaacaccaa taacgccatt taatctaagc gcatcaccaa cattttctgg cgtcagtcca

     3781 ccagctaaca taaaatgtaa gctttcgggg ctctcttgcc ttccaaccca gtcagaaatc

     3841 gagttccaat ccaaaagttc acctgtccca cctgcttctg aatcaaacaa gggaataaac

     3901 gaatgaggtt tctgtgaagc tgcactgagt agtatgttgc agtcttttgg aaatacgagt

     3961 cttttaataa ctggcaaacc gaggaactct tggtattctt gccacgactc atctccatgc

     4021 agttggacga tatcaatgcc gtaatcattg accagagcca aaacatcctc cttaggttga

     4081 ttacgaaaca cgccaaccaa gtatttcgga gtgcctgaac tatttttata tgcttttaca

     4141 agacttgaaa ttttccttgc aataaccggg tcaattgttc tctttctatt gggcacacat

     4201 ataataccca gcaagtcagc atcggaatct agagcacatt ctgcggcctc tgtgctctgc

     4261 aagccgcaaa ctttcaccaa tggaccagaa ctacctgtga aattaataac agacatactc

     4321 caagctgcct ttgtgtgctt aatcacgtat actcacgtgc tcaatagtca ccaatgccct

     4381 ccctcttggc cctctccttt tcttttttcg accgaattaa ttcttaatcg gcaaaaaaag

     4441 aaaagctccg gatcaagatt gtacgtaagg tgacaagcta tttttcaata aagaatatct

     4501 tccactactg ccatctggcg tcataactgc aaagtacaca tatattacga tgctgtctat

     4561 taaatgcttc ctatattata tatatagtaa tgtcgtttat ggtgcactct cagtacaatc

     4621 tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc

     4681 tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc

     4741 tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cga

//

 

Caution:
Product is for research use only!

 

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