LOCUS       sequence-215303-        7408 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CEN/ARS plasmid for GAL-inducible NSG2-V5-TEV-EGFP expression marked
            with URA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7408)
  AUTHORS   Costa EA, Subramanian K, Nunnari J, Weissman JS
  TITLE     Defining the physiological role of SRP in protein-targeting 
            efficiency and specificity.
  JOURNAL   Science. 2018 Jan 18. pii: science.aar3607. doi: 
            10.1126/science.aar3607.
  PUBMED    29348368
REFERENCE   2  (bases 1 to 7408)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Science. 
            2018 Jan 18. pii: science.aar3607. doi: 10.1126/science.aar3607."
COMMENT     Sequence Label: pJW1669
FEATURES             Location/Qualifiers
     source          1..7408
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        194..409
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             410..1213
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     rep_origin      complement(1344..1799)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1481..1502)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1693..1712
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1925..1947
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1939..1956
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1940..1956
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1963..1982
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1963..1981
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     2014..2030
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2014..2030
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     terminator      complement(2058..2223)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     2144..2168
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(2244..2960)
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             complement(2247..2960)
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             complement(2985..3005)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             complement(3012..3053)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     promoter        complement(3953..4394)
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     primer_bind     complement(3959..3982)
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     protein_bind    4277..4394
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     complement(4590..4606)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4591..4607)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(4635..4653)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4674..4690)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4674..4690)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4687..4709)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4698..4714
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /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(join(4722..4728,4729..4746,4747..4752))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4767..4788
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4905..4922)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5076..5664)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5156..5175)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5835..6626,6627..6695))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6458..6477
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6696..6800)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    6837..7340
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     7381..7399
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accataccac cttttcaatt catcattttt tttttattct tttttttgat ttcggtttcc
      241 ttgaaatttt tttgattcgg taatctccga acagaaggaa gaacgaagga aggagcacag
      301 acttagattg gtatatatac gcatatgtag tgttgaagaa acatgaaatt gcccagtatt
      361 cttaacccaa ctgcacagaa caaaaacctg caggaaacga agataaatca tgtcgaaagc
      421 tacatataag gaacgtgctg ctactcatcc tagtcctgtt gctgccaagc tatttaatat
      481 catgcacgaa aagcaaacaa acttgtgtgc ttcattggat gttcgtacca ccaaggaatt
      541 actggagtta gttgaagcat taggtcccaa aatttgttta ctaaaaacac atgtggatat
      601 cttgactgat ttttccatgg agggcacagt taagccgcta aaggcattat ccgccaagta
      661 caatttttta ctcttcgaag acagaaaatt tgctgacatt ggtaatacag tcaaattgca
      721 gtactctgcg ggtgtataca gaatagcaga atgggcagac attacgaatg cacacggtgt
      781 ggtgggccca ggtattgtta gcggtttgaa gcaggcggca gaagaagtaa caaaggaacc
      841 tagaggcctt ttgatgttag cagaattgtc atgcaagggc tccctatcta ctggagaata
      901 tactaagggt actgttgaca ttgcgaagag cgacaaagat tttgttatcg gctttattgc
      961 tcaaagagac atgggtggaa gagatgaagg ttacgattgg ttgattatga cacccggtgt
     1021 gggtttagat gacaagggag acgcattggg tcaacagtat agaaccgtgg atgatgtggt
     1081 ctctacagga tctgacatta ttattgttgg aagaggacta tttgcaaagg gaagggatgc
     1141 taaggtagag ggtgaacgtt acagaaaagc aggctgggaa gcatatttga gaagatgcgg
     1201 ccagcaaaac taaaaaactg tattataagt aaatgcatgt atactaaact cacaaattag
     1261 agcttcaatt taattatatc agttattacc ctatgcggtg tgaaataccg cacagatgcg
     1321 taaggagaaa ataccgcatc aggaaattgt aagcgttaat attttgttaa aattcgcgtt
     1381 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta
     1441 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc
     1501 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
     1561 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact
     1621 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
     1681 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc
     1741 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc
     1801 cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc ctcttcgcta
     1861 ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat taagttgggt aacgccaggg
     1921 ttttcccagt cacgacgttg taaaacgacg gccagtgaat tgtaatacga ctcactatag
     1981 ggcgaattgg agctccaccg cggtggcggc cgctctagaa ctagtggatc ccccgggctg
     2041 cagggtgtgg tcaataagag cgacctcata ctatacctga gaaagcaacc tgacctacag
     2101 gaaagagtta ctcaagaata agaattttcg ttttaaaacc taagagtcac tttaaaattt
     2161 gtatacactt atttttttta taacttattt aataataaaa atcataaatc ataagaaatt
     2221 cgcttattta gaagtggcgc gccttatttg tacaattcat ccataccatg ggtaatacca
     2281 gcagcagtaa caaattctaa caagaccatg tggtctctct tttcgtttgg atctttggat
     2341 aaggcagatt gagtggataa gtaatggttg tctggtaaca agactggacc atcaccaatt
     2401 ggagtatttt gttgataatg gtcagctaat tgaacagaac catcttcaat gttgtgtcta
     2461 attttgaagt taactttgat accattcttt tgtttgtcag ccatgatgta aacattgtga
     2521 gagttatagt tgtattccaa tttgtgacct aaaatgttac catcttcttt aaaatcaata
     2581 ccttttaatt cgattctatt aactaaggta tcaccttcaa acttgacttc agctctggtc
     2641 ttgtagttac cgtcatcttt gaaaaaaata gttctttctt gaacataacc ttctggcatg
     2701 gcagacttga aaaagtcatg ttgtttcata tgatctgggt atctagaaaa acattgaaca
     2761 ccataagtta aagtagtgac taaggttggc catggaactg gcaatttacc agtagtacaa
     2821 ataaatttta aggtcaattt accgtaagta gcatcacctt caccttcacc ggagacagaa
     2881 aatttgtgac cattaacatc accatctaat tcaaccaaaa ttgggacaac accagtgaat
     2941 aattcttcac ctttagacat gttaattaaa ccagcaccgt cacctccttg aaaatacaaa
     3001 ttttctgaac ccgtagaatc gagaccgagg agagggttag ggataggctt accatgagaa
     3061 tatagatatc ttcctagttt tccaaacatt agagaaccaa ggaaataaaa atccatgaaa
     3121 tataaaaatt gggaaccgtc gataatatct ttgtagcaag agatacaaca acccacccca
     3181 atgcagatgc agctcattaa catggatatt gagccgtcaa aaaacaacca cagtataata
     3241 ttcagaagcc cccatgcccc tgccgcctgc aaagaggagt tccactgtat tttcctgatc
     3301 ccaaatgtga ctcccaacat ggcattgata cttctaataa cagaggtcaa actgtcatgg
     3361 tggacagagc tcttaacaac ctctgtttga aaaatattat ccagaattgg tacaacagaa
     3421 ccaaataaca gtccttcaac accgtagcct aaccagtttg gcagaacccc attggatagc
     3481 cagttacaaa gtttgacacc cagtagaagg ggtctcgatg caaaatctgg atgtaagaga
     3541 tgattatcgt gcaagtttct ggacagttca tgatatgcga aaccgctcaa tgacaaaatt
     3601 aatacagctt gcaggtaatg aagaatattt tgacgagttt gtctggttct tgacggaaag
     3661 ttttcgtgaa tggatgatgc tgataaaatc ctttccaaat tattagaaga tactgtgttc
     3721 cttggcagcg agccgccaac cttatgggcc aagtaaaaag tcgaatcaat agtttgatct
     3781 ctactatatt tggtagaatc tatagagaca gagcgtttca attcttcgta aatttcgtta
     3841 tcttccgatc tcacgacgtc atcatcatat aatgaataaa gctgcggttt agtcagagag
     3901 cagattgatt cggtgctctt ctttggatcc ggttctcctc tattggccat ttctccttga
     3961 cgttaaagta tagaggtata ttaacaattt tttgttgata cttttatgac atttgaataa
     4021 gaagtaatac aaactgaaaa tgttgaaagt attagttaaa gtggttatgc agcttttcca
     4081 tttatatatc tgttaataga tcaaaaatca tcgcttcgct gattaattac cccagaaaca
     4141 aggctaaaaa actaatcgca ttatcatcct atggttgtta atttgattcg ttaatttgaa
     4201 ggtttgtggg gccaggttac tgccaatttt tcctcttcat aaccataaaa gctagtattg
     4261 tagaatcttt attgttcgga gcagtgcggc gcgaggcaca tctgcgtttc aggaacgcga
     4321 ccggtgaaga cgaggacgca cggaggagag tcttccgtcg gagggctgtc gcccgctcgg
     4381 cggcttctaa tccgtacttc aatatagcaa tgagcagtta agcgtattac tgaaagttcc
     4441 aaagagaagg tttttttagg ctaagataat ggggctcttt acatttccac aacatataag
     4501 taagattaga tatggatatg tatatggtgg taatgccatg taatatgatt attaaacttc
     4561 tttgcgtcca tccaagatat caagcttatc gataccgtcg acctcgaggg ggggcccggt
     4621 accagctttt gttcccttta gtgagggtta atttcgagct tggcgtaatc atggtcatag
     4681 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     4741 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     4801 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     4861 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     4921 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     4981 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     5041 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     5101 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     5161 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     5221 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     5281 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     5341 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     5401 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     5461 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     5521 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     5581 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     5641 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     5701 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     5761 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     5821 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     5881 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     5941 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     6001 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     6061 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     6121 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     6181 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     6241 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     6301 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     6361 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     6421 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     6481 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     6541 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     6601 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     6661 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     6721 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     6781 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgggtc cttttcatca
     6841 cgtgctataa aaataattat aatttaaatt ttttaatata aatatataaa ttaaaaatag
     6901 aaagtaaaaa aagaaattaa agaaaaaata gtttttgttt tccgaagatg taaaagactc
     6961 tagggggatc gccaacaaat actacctttt atcttgctct tcctgctctc aggtattaat
     7021 gccgaattgt ttcatcttgt ctgtgtagaa gaccacacac gaaaatcctg tgattttaca
     7081 ttttacttat cgttaatcga atgtatatct atttaatctg cttttcttgt ctaataaata
     7141 tatatgtaaa gtacgctttt tgttgaaatt ttttaaacct ttgtttattt ttttttcttc
     7201 attccgtaac tcttctacct tctttattta ctttctaaaa tccaaataca aaacataaaa
     7261 ataaataaac acagagtaaa ttcccaaatt attccatcat taaaagatac gaggcgcgtg
     7321 taagttacag gcaagcgatc cgtctaagaa accattatta tcatgacatt aacctataaa
     7381 aataggcgta tcacgaggcc ctttcgtc
//
