pRS425 plasmid

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  • Model: PVT4046
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pRS425 Plasmid

PVT4046     2ug

 

pRS425 plasmid informaiton

Promoter: LEU2, T7, T3
Replicator: 2 ori, ori, FI ori
Plasmid classification: yeast series, Saccharomyces cerevisiae expression vector
Plasmid size: 6849bp
Prokaryotic resistance: Amp
Screening markers: LEU2
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

 

pRS425 plasmid Description
This is a YEtype(episomal) yeast shuttle vector. It encodes the beta galactosidase alpha peptide for bluewhite color detection of inserts and also has the T7 and T3 promoters for in vitro RNA synthesis and priming sites for sequencing. It also contains the REP3 and FRT sequences necessary for high copy propagation in yeast. In S. cerevisiae, the copy number is about 20 per haploid cell. In nonselective growth, plasmids are lost through mitotic segregation at rates in the range of 4.6 +/ 0.6% of progeny per doubling. The order of the major features in this plasmid is: ori(f1) lacZ T7 promoter MCS (KpnISacI) T3 promoter lacI ori(pMB1) ampR ori(2 micron) LEU2.
pRS425-plasmid
 

pRS425 plasmid Sequence
 

LOCUS       Exported                6849 bp ds-DNA     circular SYN 12-SEP-2016

DEFINITION  synthetic circular DNA

ACCESSION   .

VERSION     .

KEYWORDS    Untitled 7

SOURCE      synthetic DNA construct

  ORGANISM  synthetic DNA construct

REFERENCE   1  (bases 1 to 6849)

  AUTHORS   .

  TITLE     Direct Submission

  JOURNAL   Exported Monday, September 12, 2016 from SnapGene Viewer 3.1.4

FEATURES             Location/Qualifiers

     source          1..6849

                     /organism="synthetic DNA construct"

                     /mol_type="other DNA"

     CDS             complement(663..1757)

                     /codon_start=1

                     /gene="S. cerevisiae LEU2"

                     /product="3-isopropylmalate dehydrogenase, required for 

                     leucine biosynthesis"

                     /note="LEU2"

                     /note="yeast auxotrophic marker"

                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL

                     IGGAAIDATGVPLPDEALEASKKVDAVLLGAVAGPKWGTGSVRPEQGLLKIRKELQLYA

                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT

                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANLLASSRLWRKTVEETIKNEFPTLKVQHQ

                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF

                     GLYEPCHGSAPDLPKNKVDPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG

                     DLGGSNSTTEVGDAVAEEVKKILA"

     promoter        complement(1770..2174)

                     /gene="S. cerevisiae LEU2"

                     /note="LEU2 promoter"

     rep_origin      complement(2474..2929)

                     /direction=LEFT

                     /note="f1 ori"

                     /note="f1 bacteriophage origin of replication; arrow 

                     indicates direction of (+) strand synthesis"

     CDS             complement(2712..3290)

                     /codon_start=1

                     /gene="lacZ fragment"

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

                     /note="lacZ-alpha"

                     /translation="MTMITPSAQLTLTKGNKSWSSTAVAAALELVDPPGCRNSISSLSI

                     PSTSRGGPVPNSPYSESYYARSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEAR

                     TDRPSQQLRSLNGEWRDAPCSGALSAAGVVVTRSVTATLASALAPAPFAFFPSFLATFA

                     GFPRQALNRGLPLGFRFSALRHLDPKKLD"

     primer_bind     3074..3090

                     /note="M13 fwd"

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

                     variants"

     promoter        3100..3118

                     /note="T7 promoter"

                     /note="promoter for bacteriophage T7 RNA polymerase"

     misc_feature    3127..3234

                     /note="MCS"

                     /note="pBluescript multiple cloning site"

     primer_bind     3144..3160

                     /note="KS primer"

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

                     variants"

     primer_bind     complement(3194..3210)

                     /note="SK primer"

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

                     variants"

     promoter        complement(3247..3265)

                     /note="T3 promoter"

                     /note="promoter for bacteriophage T3 RNA polymerase"

     primer_bind     complement(3286..3302)

                     /note="M13 rev"

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

                     variants"

     protein_bind    3310..3326

                     /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)."

     promoter        complement(3334..3364)

                     /note="lac promoter"

                     /note="promoter for the E. coli lac operon"

     rep_origin      complement(3688..4276)

                     /direction=LEFT

                     /note="ori"

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

                     replication"

     CDS             complement(4447..5307)

                     /codon_start=1

                     /gene="bla"

                     /product="beta-lactamase"

                     /note="AmpR"

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

                     related antibiotics"

                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI

                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS

                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW

                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA

                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS

                     LIKHW"

     promoter        complement(5308..5412)

                     /gene="bla"

                     /note="AmpR promoter"

     rep_origin      5439..6781

                     /note="2u ori"

                     /note="yeast 2u plasmid origin of replication"

     protein_bind    complement(6402..6449)

                     /bound_moiety="FLP recombinase from the Saccharomyces 

                     cerevisiae 2u plasmid"

                     /note="FRT"

                     /note="FLP-mediated recombination occurs in the 8-bp core 

                     sequence TCTAGAAA (Turan and Bode, 2011)."

ORIGIN

        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca

       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg

      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc

      181 accatatcga ctacgtcgta aggccgtttc tgacagagta aaattcttga gggaactttc

      241 accattatgg gaaatgcttc aagaaggtat tgacttaaac tccatcaaat ggtcaggtca

      301 ttgagtgttt tttatttgtt gtattttttt ttttttagag aaaatcctcc aatatcaaat

      361 taggaatcgt agtttcatga ttttctgtta cacctaactt tttgtgtggt gccctcctcc

      421 ttgtcaatat taatgttaaa gtgcaattct ttttccttat cacgttgagc cattagtatc

      481 aatttgctta cctgtattcc tttactatcc tcctttttct ccttcttgat aaatgtatgt

      541 agattgcgta tatagtttcg tctaccctat gaacatattc cattttgtaa tttcgtgtcg

      601 tttctattat gaatttcatt tataaagttt atgtacaaat atcataaaaa aagagaatct

      661 ttttaagcaa ggattttctt aacttcttcg gcgacagcat caccgacttc ggtggtactg

      721 ttggaaccac ctaaatcacc agttctgata cctgcatcca aaaccttttt aactgcatct

      781 tcaatggcct taccttcttc aggcaagttc aatgacaatt tcaacatcat tgcagcagac

      841 aagatagtgg cgatagggtc aaccttattc tttggcaaat ctggagcaga accgtggcat

      901 ggttcgtaca aaccaaatgc ggtgttcttg tctggcaaag aggccaagga cgcagatggc

      961 aacaaaccca aggaacctgg gataacggag gcttcatcgg agatgatatc accaaacatg

     1021 ttgctggtga ttataatacc atttaggtgg gttgggttct taactaggat catggcggca

     1081 gaatcaatca attgatgttg aaccttcaat gtagggaatt cgttcttgat ggtttcctcc

     1141 acagtttttc tccataatct tgaagaggcc aaaagattag ctttatccaa ggaccaaata

     1201 ggcaatggtg gctcatgttg tagggccatg aaagcggcca ttcttgtgat tctttgcact

     1261 tctggaacgg tgtattgttc actatcccaa gcgacaccat caccatcgtc ttcctttctc

     1321 ttaccaaagt aaatacctcc cactaattct ctgacaacaa cgaagtcagt acctttagca

     1381 aattgtggct tgattggaga taagtctaaa agagagtcgg atgcaaagtt acatggtctt

     1441 aagttggcgt acaattgaag ttctttacgg atttttagta aaccttgttc aggtctaaca

     1501 ctaccggtac cccatttagg accagccaca gcacctaaca aaacggcatc aaccttcttg

     1561 gaggcttcca gcgcctcatc tggaagtggg acacctgtag catcgatagc agcaccacca

     1621 attaaatgat tttcgaaatc gaacttgaca ttggaacgaa catcagaaat agctttaaga

     1681 accttaatgg cttcggctgt gatttcttga ccaacgtggt cacctggcaa aacgacgatc

     1741 ttcttagggg cagacatagg ggcagacatt agaatggtat atccttgaaa tatatatata

     1801 tattgctgaa atgtaaaagg taagaaaagt tagaaagtaa gacgattgct aaccacctat

     1861 tggaaaaaac aataggtcct taaataatat tgtcaacttc aagtattgtg atgcaagcat

     1921 ttagtcatga acgcttctct attctatatg aaaagccggt tccggcctct cacctttcct

     1981 ttttctccca atttttcagt tgaaaaaggt atatgcgtca ggcgacctct gaaattaaca

     2041 aaaaatttcc agtcatcgaa tttgattctg tgcgatagcg cccctgtgtg ttctcgttat

     2101 gttgaggaaa aaaataatgg ttgctaagag attcgaactc ttgcatctta cgatacctga

     2161 gtattcccac agttaactgc ggtcaagata tttcttgaat caggcgcctt agaccgctcg

     2221 gccaaacaac caattacttg ttgagaaata gagtataatt atcctataaa tataacgttt

     2281 ttgaacacac atgaacaagg aagtacagga caattgattt tgaagagaat gtggattttg

     2341 atgtaattgt tgggattcca tttttaataa ggcaataata ttaggtatgt ggatatacta

     2401 gaagttctcc tcgaccgtcg atatgcggtg tgaaataccg cacagatgcg taaggagaaa

     2461 ataccgcatc aggaaattgt aaacgttaat attttgttaa aattcgcgtt aaatttttgt

     2521 taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa

     2581 gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag

     2641 aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt

     2701 gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac

     2761 cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag

     2821 gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg

     2881 cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc gcgccattcg

     2941 ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc

     3001 cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc

     3061 cagtcacgac gttgtaaaac gacggccagt gagcgcgcgt aatacgactc actatagggc

     3121 gaattgggta ccgggccccc cctcgaggtc gacggtatcg ataagcttga tatcgaattc

     3181 ctgcagcccg ggggatccac tagttctaga gcggccgcca ccgcggtgga gctccagctt

     3241 ttgttccctt tagtgagggt taattgcgcg cttggcgtaa tcatggtcat agctgtttcc

     3301 tgtgtgaaat tgttatccgc tcacaattcc acacaacata ggagccggaa gcataaagtg

     3361 taaagcctgg ggtgcctaat gagtgaggta actcacatta attgcgttgc gctcactgcc

     3421 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg

     3481 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc

     3541 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac

     3601 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa

     3661 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca

     3721 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc

     3781 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata

     3841 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta

     3901 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca

     3961 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga

     4021 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg

     4081 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg

     4141 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg

     4201 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag

     4261 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa

     4321 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat

     4381 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc

     4441 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc

     4501 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc

     4561 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc

     4621 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc

     4681 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt

     4741 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc

     4801 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa

     4861 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt

     4921 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg

     4981 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc

     5041 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa

     5101 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt

     5161 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt

     5221 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag

     5281 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta

     5341 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat

     5401 aggggttccg cgcacatttc cccgaaaagt gccacctgaa cgaagcatct gtgcttcatt

     5461 ttgtagaaca aaaatgcaac gcgagagcgc taatttttca aacaaagaat ctgagctgca

     5521 tttttacaga acagaaatgc aacgcgaaag cgctatttta ccaacgaaga atctgtgctt

     5581 catttttgta aaacaaaaat gcaacgcgag agcgctaatt tttcaaacaa agaatctgag

     5641 ctgcattttt acagaacaga aatgcaacgc gagagcgcta ttttaccaac aaagaatcta

     5701 tacttctttt ttgttctaca aaaatgcatc ccgagagcgc tatttttcta acaaagcatc

     5761 ttagattact ttttttctcc tttgtgcgct ctataatgca gtctcttgat aactttttgc

     5821 actgtaggtc cgttaaggtt agaagaaggc tactttggtg tctattttct cttccataaa

     5881 aaaagcctga ctccacttcc cgcgtttact gattactagc gaagctgcgg gtgcattttt

     5941 tcaagataaa ggcatccccg attatattct ataccgatgt ggattgcgca tactttgtga

     6001 acagaaagtg atagcgttga tgattcttca ttggtcagaa aattatgaac ggtttcttct

     6061 attttgtctc tatatactac gtataggaaa tgtttacatt ttcgtattgt tttcgattca

     6121 ctctatgaat agttcttact acaatttttt tgtctaaaga gtaatactag agataaacat

     6181 aaaaaatgta gaggtcgagt ttagatgcaa gttcaaggag cgaaaggtgg atgggtaggt

     6241 tatataggga tatagcacag agatatatag caaagagata cttttgagca atgtttgtgg

     6301 aagcggtatt cgcaatattt tagtagctcg ttacagtccg gtgcgttttt ggttttttga

     6361 aagtgcgtct tcagagcgct tttggttttc aaaagcgctc tgaagttcct atactttcta

     6421 gagaatagga acttcggaat aggaacttca aagcgtttcc gaaaacgagc gcttccgaaa

     6481 atgcaacgcg agctgcgcac atacagctca ctgttcacgt cgcacctata tctgcgtgtt

     6541 gcctgtatat atatatacat gagaagaacg gcatagtgcg tgtttatgct taaatgcgta

     6601 cttatatgcg tctatttatg taggatgaaa ggtagtctag tacctcctgt gatattatcc

     6661 cattccatgc ggggtatcgt atgcttcctt cagcactacc ctttagctgt tctatatgct

     6721 gccactcctc aattggatta gtctcatcct tcaatgctat catttccttt gatattggat

     6781 catactaaga aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc

     6841 cctttcgtc

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