LOCUS       sequence-210833-        7182 bp    DNA     circular SYN 15-APR-2025
DEFINITION  CEN/ARS plasmid for GAL-inducible ORM1-V5-TEV-EGFP expression marked
            with URA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7182)
  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 7182)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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: pJW1671
FEATURES             Location/Qualifiers
     source          1..7182
                     /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        190..410
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             411..1214
                     /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(1343..1798)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1480..1501)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1692..1711
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1928..1950
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1942..1959
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1943..1959
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1966..1985
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1966..1984
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     2017..2033
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2017..2033
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     terminator      complement(2061..2226)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     2147..2171
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(2247..2963)
                     /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(2250..2963)
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             complement(2988..3008)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             complement(3015..3056)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     promoter        complement(3725..4166)
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     primer_bind     complement(3731..3754)
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     protein_bind    4049..4166
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     complement(4362..4378)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4363..4379)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(4408..4426)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4447..4463)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4447..4463)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4460..4482)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4471..4487
                     /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(4495..4501,4502..4519,4520..4525))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(4678..4695)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4849..5437)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5608..6399,6400..6468))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFFHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6231..6250
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6469..6573)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    6610..7113
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     7155..7173
                     /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 accacgcttt tcaattcaat tcatcatttt ttttttattc ttttttttga tttcggtttc
      241 tttgaaattt ttttgattcg gtaatctccg aacagaagga agaacgaagg aaggagcaca
      301 gacttagatt ggtatatata cgcatatgta gtgttgaaga aacatgaaat tgcccagtat
      361 tcttaaccca actgcacaga acaaaaacct gcaggaaacg aagataaatc atgtcgaaag
      421 ctacatataa ggaacgtgct gctactcatc ctagtcctgt tgctgccaag ctatttaata
      481 tcatgcacga aaagcaaaca aacttgtgtg cttcattgga tgttcgtacc accaaggaat
      541 tactggagtt agttgaagca ttaggtccca aaatttgttt actaaaaaca catgtggata
      601 tcttgactga tttttccatg gagggcacag ttaagccgct aaaggcatta tccgccaagt
      661 acaatttttt actcttcgaa gacagaaaat ttgctgacat tggtaataca gtcaaattgc
      721 agtactctgc gggtgtatac agaatagcag aatgggcaga cattacgaat gcacacggtg
      781 tggtgggccc aggtattgtt agcggtttga agcaggcggc agaagaagta acaaaggaac
      841 ctagaggcct tttgatgtta gcagaattgt catgcaaggg ctccctatct actggagaat
      901 atactaaggg tactgttgac attgcgaaga gcgacaaaga ttttgttatc ggctttattg
      961 ctcaaagaga catgggtgga agagatgaag gttacgattg gttgattatg acacccggtg
     1021 tgggtttaga tgacaaggga gacgcattgg gtcaacagta tagaaccgtg gatgatgtgg
     1081 tctctacagg atctgacatt attattgttg gaagaggact atttgcaaag ggaagggatg
     1141 ctaaggtaga gggtgaacgt tacagaaaag caggctggga agcatatttg agaagatgcg
     1201 gccagcaaaa ctaaaaaact gtattataag taaatgcatg tatactaaac tcacaaatta
     1261 gagcttcaat ttaattatat cagttattac cctgcggtgt gaaataccgc acagatgcgt
     1321 aaggagaaaa taccgcatca ggaaattgta aacgttaata ttttgttaaa attcgcgtta
     1381 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     1441 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     1501 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     1561 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     1621 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     1681 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg
     1741 gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcg
     1801 cgccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg
     1861 ctattacgcc agctggcgaa ggggggatgt gctgcaaggc gattaagttg ggtaacgcca
     1921 gggttttccc agtcacgacg ttgtaaaacg acggccagtg aattgtaata cgactcacta
     1981 tagggcgaat tggagctcca ccgcggtggc ggccgctcta gaactagtgg atcccccggg
     2041 ctgcagggtg tggtcaataa gagcgacctc atactatacc tgagaaagca acctgaccta
     2101 caggaaagag ttactcaaga ataagaattt tcgttttaaa acctaagagt cactttaaaa
     2161 tttgtataca cttatttttt ttataactta tttaataata aaaatcataa atcataagaa
     2221 attcgcttat ttagaagtgg cgcgccttat ttgtacaatt catccatacc atgggtaata
     2281 ccagcagcag taacaaattc taacaagacc atgtggtctc tcttttcgtt tggatctttg
     2341 gataaggcag attgagtgga taagtaatgg ttgtctggta acaagactgg accatcacca
     2401 attggagtat tttgttgata atggtcagct aattgaacag aaccatcttc aatgttgtgt
     2461 ctaattttga agttaacttt gataccattc ttttgtttgt cagccatgat gtaaacattg
     2521 tgagagttat agttgtattc caatttgtga cctaaaatgt taccatcttc tttaaaatca
     2581 atacctttta attcgattct attaactaag gtatcacctt caaacttgac ttcagctctg
     2641 gtcttgtagt taccgtcatc tttgaaaaaa atagttcttt cttgaacata accttctggc
     2701 atggcagact tgaaaaagtc atgttgtttc atatgatctg ggtatctaga aaaacattga
     2761 acaccataag ttaaagtagt gactaaggtt ggccatggaa ctggcaattt accagtagta
     2821 caaataaatt ttaaggtcaa tttaccgtaa gtagcatcac cttcaccttc accggagaca
     2881 gaaaatttgt gaccattaac atcaccatct aattcaacca aaattgggac aacaccagtg
     2941 aataattctt cacctttaga catgttaatt aaaccagcac cgtcacctcc ttgaaaatac
     3001 aaattttctg aacccgtaga atcgagaccg aggagagggt tagggatagg cttaccacta
     3061 atttgggcgc gacctgtgat acctgggata gaaatcctta atctatgagt aacaggtaac
     3121 tttgggacaa cagcaccaaa ggttgtcaaa aaacaattcc atgaaaacaa tttcaaatca
     3181 tagtgagcat aatgagtact aactaagaat agggcgatcg ggacactaat caaaaatttt
     3241 cttgaaggag tatataaagt ctcgtcgtca atttgttccc acatcgtcaa gttgtcataa
     3301 gcaccaccat tgaaatcgaa aggggtaccc ttaatcagat ggaacatgac ataggaccca
     3361 ataacatatg tcatattagt cagagtccac gaccattctg tggtaacacc aggaaataaa
     3421 ttataaaata gtttcagcag tatgataatg accacatgaa taatccaagc gccgcgttgg
     3481 tctacccaag tagcattcat atttggtaga agttgctggt cattttcatc ttcaaaagtc
     3541 tccggttcca catgtgaaat tatgctggaa gaacgccttc ttctatgatc tttcacaggt
     3601 tccgtcgttt taattgatga agatgcactg cctgcagaag gaaacggctt aaggtccgtt
     3661 tgatttcgac tatacgaggt agaagccgcc tcagcagttc cttgataatc taattcggtc
     3721 atttctcctt gacgttaaag tatagaggta tattaacaat tttttgttga tacttttatg
     3781 acatttgaat aagaagtaat acaaactgaa aatgttgaaa gtattagtta aagtggttat
     3841 gcagcttttc catttatata tctgttaata gatcaaaaat catcgcttcg ctgattaatt
     3901 accccagaaa caaggctaaa aaactaatcg cattatcatc ctatggttgt taatttgatt
     3961 cgttaatttg aaggtttgtg gggccaggtt actgccaatt tttcctcttc ataaccataa
     4021 aagctagtat tgtagaatct ttattgttcg gagcagtgcg gcgcgaggca catctgcgtt
     4081 tcaggaacgc gaccggtgaa gacgaggacg cacggaggag agtcttccgt cggagggctg
     4141 tcgcccgctc ggcggcttct aatccgtact tcaatatagc aatgagcagt taagcgtatt
     4201 actgaaagtt ccaaagagaa ggttttttta ggctaagata atggggctct ttacatttcc
     4261 acaacatata agtaagatta gatatggata tgtatatggt ggtaatgcca tgtaatatga
     4321 ttattaaact tctttgcgtc catccaagat atcaagctta tcgataccgt cgacctcgag
     4381 ggggggcccg gtacccagct tttgttccct ttagtgaggg ttaattccga gcttggcgta
     4441 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     4501 aggagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgaggt aactcacatt
     4561 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     4621 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     4681 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     4741 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     4801 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     4861 cggcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     4921 aggactataa agataccagg cgttcccccc tggaagctcc ctcgtgcgct ctcctgttcc
     4981 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     5041 tcaatgctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     5101 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     5161 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     5221 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     5281 cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     5341 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     5401 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     5461 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     5521 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     5581 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     5641 agcgatctgt ctatttcgtt catccatagt tgcctgactg cccgtcgtgt agataactac
     5701 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     5761 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     5821 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     5881 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     5941 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     6001 atgatccccc atgttgtgaa aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     6061 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     6121 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     6181 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     6241 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     6301 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     6361 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     6421 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     6481 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     6541 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctgg
     6601 gtccttttca tcacgtgcta taaaaataat tataatttaa attttttaat ataaatatat
     6661 aaattaaaaa tagaaagtaa aaaaagaaat taaagaaaaa atagtttttg ttttccgaag
     6721 atgtaaaaga ctctaggggg atcgccaaca aatactacct tttatcttgc tcttcctgct
     6781 ctcaggtatt aatgccgaat tgtttcatct tgtctgtgta gaagaccaca cacgaaaatc
     6841 ctgtgatttt acattttact tatcgttaat cgaatgtata tctatttaat ctgcttttct
     6901 tgtctaataa atatatatgt aaagtacgct ttttgttgaa attttttaaa cctttgttta
     6961 tttttttttc ttcattccgt aactcttcta ccttctttat ttactttcta aaatccaaat
     7021 acaaaacata aaaataaata aacacagagt aaattcccaa attattccat cattaaaaga
     7081 tacgaggcgc gtgtaagtta caggcaagcg atccgtccta agaaaccatt attatcatga
     7141 cattaaccta taaaaatagg cgtatcacga ggccctttcg tc
//
