LOCUS       sequence-245407-        7973 bp    DNA     circular SYN 14-APR-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 7973)
  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 7973)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 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: DRD1-Tango
FEATURES             Location/Qualifiers
     source          1..7973
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(2531..3145,3146..3532)
                     /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(2598..2617)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(3564..3581)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3570..3794
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3840..4268
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3927..3946)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4137..4158
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4277..4297)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4282..4611
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4462..4597
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4524..4543
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4678..5472
                     /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(4732..4751)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5342..5361
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5646..5767
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5683..5702)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5737..5756
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5816..5832)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5816..5832)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5829..5851)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5840..5856
                     /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(5864..5870,5871..5888,5889..5894))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5909..5930
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6047..6064)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6218..6806)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6298..6317)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6977..7768,7769..7837))
                     /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     7600..7619
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7838..7942)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttggta ccgagctcgg atccactagt ccagtgtggt ggaattctgc
      961 agatatccag cacagtggcg gccgcgccac catgaagacg atcatcgccc tgagctacat
     1021 cttctgcctg gtattcgccg actacaagga cgatgatgac gccagcatcg atatgcgcac
     1081 attgaatact tcagcgatgg acggaacggg cctcgtggtc gaacgggact ttagcgttag
     1141 aattcttacc gcttgcttcc tgtcactgct tatcctgtct actctcctgg gtaatacact
     1201 tgtgtgcgcc gccgtgatca gattccggca cttgcggtcc aaagtgacta acttttttgt
     1261 catctcactg gccgtctctg atctgctggt ggctgtcctc gtgatgccat ggaaagcagt
     1321 ggccgagatc gccgggttct ggcccttcgg gtcattttgc aatatctggg tggcattcga
     1381 catcatgtgt agcacggcct caattctcaa tctctgtgtc attagtgtcg ataggtattg
     1441 ggccatcagc tctcccttcc gatacgagcg gaagatgacc ccaaaggccg cctttatact
     1501 gatatcagtc gcctggaccc tcagcgtgct gatttctttc atccctgtgc aattgtcctg
     1561 gcacaaggct aaaccaacga gtcccagcga cggaaacgcc actagtctgg ccgagactat
     1621 cgacaattgt gacagtagcc tgtccaggac ctacgctatc agcagttcag tgatctcctt
     1681 ttacattcct gtcgcgatta tgatcgtgac ctacacaagg atctatagaa tcgcacagaa
     1741 gcaaatccgg aggatcgcgg ctttggagag agccgctgtg cacgcaaaga attgtcagac
     1801 gacgaccggc aacggtaaac ccgtcgagtg ttcccagcca gagtccagct tcaaaatgtc
     1861 atttaagcgc gaaactaagg ttctgaagac cctctctgtg attatgggcg tgtttgtgtg
     1921 ttgctggctc cccttcttca ttttgaattg tatactgcct ttctgcggca gtggcgagac
     1981 tcagcctttc tgtatagact caaacacttt tgacgtcttc gtttggttcg gttgggccaa
     2041 ttcctccctt aatcctatta tatacgcatt taatgctgat tttcgcaaag cattctccac
     2101 cctgctgggc tgttaccggc tttgtcccgc caccaataac gcaattgaga ccgtatctat
     2161 taacaataac ggcgcagcta tgttctcctc tcaccatgag ccaagaggtt caattagcaa
     2221 agagtgtaat cttgtttact tgatacctca cgcagttggg agttcagaag accttaagaa
     2281 agaagaagcc gccggcatcg ctaggcccct cgaaaaactg agccctgcac tttctgtgat
     2341 cctggattat gacactgatg tgagcctgga gaaaattcaa ccaataacac agaacggtca
     2401 acatccaact atcgataccg gtggacgcac cccacccagc ctgggtcccc aagatgagtc
     2461 ctgcaccacc gccagctcct ccctggccaa ggacacttca tcgaccggtg agaacctgta
     2521 cttccagcta agattagata aaagtaaagt gattaacagc gcattagagc tgcttaatga
     2581 ggtcggaatc gaaggtttaa caacccgtaa actcgcccag aagctaggtg tagagcagcc
     2641 tacattgtat tggcatgtaa aaaataagcg ggctttgctc gacgccttag ccattgagat
     2701 gttagatagg caccatactc acttttgccc tttagaaggg gaaagctggc aagatttttt
     2761 acgtaataac gctaaaagtt ttagatgtgc tttactaagt catcgcgatg gagcaaaagt
     2821 acatttaggt acacggccta cagaaaaaca gtatgaaact ctcgaaaatc aattagcctt
     2881 tttatgccaa caaggttttt cactagagaa tgcattatat gcactcagcg ctgtggggca
     2941 ttttacttta ggttgcgtat tggaagatca agagcatcaa gtcgctaaag aagaaaggga
     3001 aacacctact actgatagta tgccgccatt attacgacaa gctatcgaat tatttgatca
     3061 ccaaggtgca gagccagcct tcttattcgg ccttgaattg atcatatgcg gattagaaaa
     3121 acaacttaaa tgtgaaagtg ggtccgcgta cagccgcgcg cgtacgaaaa acaattacgg
     3181 gtctaccatc gagggcctgc tcgatctccc ggacgacgac gcccccgaag aggcggggct
     3241 ggcggctccg cgcctgtcct ttctccccgc gggacacacg cgcagactgt cgacggcccc
     3301 cccgaccgat gtcagcctgg gggacgagct ccacttagac ggcgaggacg tggcgatggc
     3361 gcatgccgac gcgctagacg atttcgatct ggacatgttg ggggacgggg attccccggg
     3421 tccgggattt accccccacg actccgcccc ctacggcgct ctggatatgg ccgacttcga
     3481 gtttgagcag atgtttaccg atgcccttgg aattgacgag tacggtgggt agtctagagg
     3541 gcccgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt
     3601 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
     3661 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
     3721 ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc
     3781 ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca
     3841 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     3901 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     3961 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     4021 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     4081 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     4141 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     4201 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     4261 cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     4321 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     4381 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     4441 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     4501 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag
     4561 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg
     4621 ggagcttgta tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg
     4681 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     4741 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     4801 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag
     4861 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     4921 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     4981 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     5041 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     5101 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     5161 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc
     5221 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     5281 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     5341 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     5401 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     5461 gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc
     5521 catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     5581 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     5641 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     5701 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     5761 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     5821 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     5881 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     5941 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     6001 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     6061 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     6121 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     6181 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     6241 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     6301 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     6361 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     6421 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     6481 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     6541 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     6601 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     6661 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     6721 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     6781 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     6841 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     6901 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     6961 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     7021 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     7081 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     7141 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     7201 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     7261 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     7321 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     7381 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     7441 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     7501 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     7561 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     7621 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     7681 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     7741 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     7801 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     7861 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     7921 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtc
//
