LOCUS       sequence-247685-        7682 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of G protein-coupled receptors for PRESTO-Tango: parallel
            receptorome expression and screening via transcriptional output, 
            with transcriptional activation following arrestin translocation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7682)
  AUTHORS   Kroeze WK, Sassano MF, Huang XP, Lansu K, McCorvy JD, Giguere PM, 
            Sciaky N, Roth BL
  TITLE     PRESTO-Tango as an open-source resource for interrogation of the 
            druggable human GPCRome.
  JOURNAL   Nat Struct Mol Biol. 2015 May;22(5):362-9. doi: 10.1038/nsmb.3014. 
            Epub 2015 Apr 20.
  PUBMED    25895059
REFERENCE   2  (bases 1 to 7682)
  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"; doi: "10.1038/nsmb.3014";
            journalName: "Nat Struct Mol Biol"; date: "2015-05"; volume: "22"; 
            issue: "5"; pages: "362-9"
COMMENT     Sequence Label: GAL1-Tango
FEATURES             Location/Qualifiers
     source          1..7682
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(46..65)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        237..616
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        617..820
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     771..791
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     865..884
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        865..883
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(2242..2856,2857..3243)
                     /codon_start=1
                     /product="tetracycline-controlled transactivator, 
                     comprising a fusion of the tetracycline repressor TetR with
                     the C-terminal activation domain of herpes simplex virus 
                     VP16"
                     /label=tTA
                     /note="In the Tet-Off(R) system, tTA binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the absence of tetracycline or doxycycline."
                     /translation="RLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHVKN
                     KRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTRPT
                     EKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTTDS
                     MPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGSAYSRARTKNNYGSTIEG
                     LLDLPDDDAPEEAGLAAPRLSFLPAGHTRRLSTAPPTDVSLGDELHLDGEDVAMAHADA
                     LDDFDLDMLGDGDSPGPGFTPHDSAPYGALDMADFEFEQMFTDALGIDEYGG"
     primer_bind     complement(2309..2328)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(3275..3292)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3281..3505
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3551..3979
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3638..3657)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3848..3869
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3988..4008)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3993..4322
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4173..4308
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4235..4254
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4389..5183
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4443..4462)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5053..5072
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5357..5478
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5394..5413)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5448..5467
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5527..5543)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5527..5543)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5540..5562)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5551..5567
                     /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(5575..5581,5582..5599,5600..5605))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5620..5641
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5758..5775)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5929..6517)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6009..6028)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6688..7479,7480..7548))
                     /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     7311..7330
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7549..7653)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga
       61 tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg
      121 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct
      181 gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca
      241 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata
      301 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
      361 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
      421 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
      481 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
      541 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
      601 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
      661 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
      721 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
      781 cggtaggcgt gtacggtggg aggtctatat aagcagagct ctctggctaa ctagagaacc
      841 cactgcttac tggcttatcg aaattaatac gactcactat agggagaccc aagctggcta
      901 gcgtttaaac ttaagcttgg taccgagctc ggatccacta gtccagtgtg gtggaattct
      961 gcagatatcc agcacagtgg cggccgcgcc accatgaaga cgatcatcgc cctgagctac
     1021 atcttctgcc tggtattcgc cgactacaag gacgatgatg acgccagcat cgatatggaa
     1081 ctggcggtcg gcaatcttag cgaaggcaac gcttcctggc cagagccccc agcacccgag
     1141 cctgggccgc tgtttggcat tggcgttgaa aattttgtga ctctggtcgt ctttgggctg
     1201 atattcgctc tgggggtgct tggaaactct ctggtcatta cagtgcttgc taggagtaaa
     1261 ccagggaagc caagatctac tacaaacctc tttatactta acctgtccat cgcagatctg
     1321 gcttatctgc tgttctgcat ccctttccag gccacagtct acgcccttcc gacttgggtc
     1381 ctgggggctt tcatctgcaa gtttattcac tattttttca cggtctcaat gttggtttcc
     1441 atttttaccc tcgctgcaat gagcgtagat cgctacgtag ctatcgtgca ctctaggaga
     1501 tctagctctc tgcgcgtatc ccggaatgct cttctgggtg tcggctgcat ttgggcactc
     1561 agcattgcga tggcttcacc tgtggcctat caccagggcc tgtttcaccc acgggcctct
     1621 aatcaaacat tctgttggga acagtggcca gaccccaggc acaagaaagc atatgtggtt
     1681 tgcacttttg tgtttggcta tctcctgcca ctcctgctca tctgtttctg ttatgctaaa
     1741 gtattgaatc acttgcacaa aaagctcaaa aatatgtcca agaagtctga agcatccaag
     1801 aagaaaacag cacagacggt gctggttgta gtggtcgtct tcggaatatc atggctgcct
     1861 caccacatca tacacctctg ggctgagttc ggagtgttcc cgctcactcc cgcttctttt
     1921 ctctttagaa tcacagccca ctgcctcgcg tactcaaact cttcagtgaa ccccataatc
     1981 tatgcttttc tgagcgagaa ctttagaaag gcctacaaac aggttttcaa gtgtcacata
     2041 cgcaaagact ctcaccttag tgatactaag gagtccaagt ccaggattga tacacctcct
     2101 tcaacaaact gcacccacgt aatcgatacc ggtggacgca ccccacccag cctgggtccc
     2161 caagatgagt cctgcaccac cgccagctcc tccctggcca aggacacttc atcgaccggt
     2221 gagaacctgt acttccagct aagattagat aaaagtaaag tgattaacag cgcattagag
     2281 ctgcttaatg aggtcggaat cgaaggttta acaacccgta aactcgccca gaagctaggt
     2341 gtagagcagc ctacattgta ttggcatgta aaaaataagc gggctttgct cgacgcctta
     2401 gccattgaga tgttagatag gcaccatact cacttttgcc ctttagaagg ggaaagctgg
     2461 caagattttt tacgtaataa cgctaaaagt tttagatgtg ctttactaag tcatcgcgat
     2521 ggagcaaaag tacatttagg tacacggcct acagaaaaac agtatgaaac tctcgaaaat
     2581 caattagcct ttttatgcca acaaggtttt tcactagaga atgcattata tgcactcagc
     2641 gctgtggggc attttacttt aggttgcgta ttggaagatc aagagcatca agtcgctaaa
     2701 gaagaaaggg aaacacctac tactgatagt atgccgccat tattacgaca agctatcgaa
     2761 ttatttgatc accaaggtgc agagccagcc ttcttattcg gccttgaatt gatcatatgc
     2821 ggattagaaa aacaacttaa atgtgaaagt gggtccgcgt acagccgcgc gcgtacgaaa
     2881 aacaattacg ggtctaccat cgagggcctg ctcgatctcc cggacgacga cgcccccgaa
     2941 gaggcggggc tggcggctcc gcgcctgtcc tttctccccg cgggacacac gcgcagactg
     3001 tcgacggccc ccccgaccga tgtcagcctg ggggacgagc tccacttaga cggcgaggac
     3061 gtggcgatgg cgcatgccga cgcgctagac gatttcgatc tggacatgtt gggggacggg
     3121 gattccccgg gtccgggatt taccccccac gactccgccc cctacggcgc tctggatatg
     3181 gccgacttcg agtttgagca gatgtttacc gatgcccttg gaattgacga gtacggtggg
     3241 tagtctagag ggcccgttta aacccgctga tcagcctcga ctgtgccttc tagttgccag
     3301 ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact
     3361 gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt
     3421 ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat
     3481 gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg
     3541 gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     3601 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     3661 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     3721 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     3781 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     3841 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     3901 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     3961 caaaaattta acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc
     4021 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt
     4081 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     4141 cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg
     4201 cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct
     4261 ctgcctctga gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca
     4321 aaaagctccc gggagcttgt atatccattt tcggatctga tcaagagaca ggatgaggat
     4381 cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga
     4441 ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc
     4501 ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga
     4561 atgaactgca ggacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg
     4621 cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc
     4681 cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg
     4741 atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga
     4801 aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc
     4861 tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca
     4921 tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg
     4981 tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct
     5041 atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg
     5101 accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc
     5161 gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac
     5221 gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     5281 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     5341 gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     5401 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     5461 actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta
     5521 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     5581 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     5641 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     5701 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     5761 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     5821 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     5881 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     5941 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     6001 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     6061 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     6121 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     6181 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     6241 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     6301 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     6361 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     6421 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     6481 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     6541 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     6601 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     6661 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     6721 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     6781 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     6841 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     6901 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     6961 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     7021 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     7081 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     7141 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     7201 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     7261 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     7321 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     7381 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     7441 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     7501 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     7561 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     7621 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     7681 cg
//
