LOCUS       sequence-203070-        7706 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 7706)
  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 7706)
  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: GPRC5A-Tango
FEATURES             Location/Qualifiers
     source          1..7706
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(73..92)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        264..643
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        644..847
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     798..818
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     892..911
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        892..910
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(2293..2907,2908..3294)
                     /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(2360..2379)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(3326..3343)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3332..3556
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3602..4030
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3689..3708)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3899..3920
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4039..4059)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4044..4373
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4224..4359
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4286..4305
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4440..5234
                     /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(4494..4513)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5104..5123
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5408..5529
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5445..5464)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5499..5518
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5578..5594)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5578..5594)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5591..5613)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5602..5618
                     /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(5626..5632,5633..5650,5651..5656))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5671..5692
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5809..5826)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5980..6568)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6060..6079)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6739..7530,7531..7599))
                     /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     7362..7381
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7600..7704)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg
       61 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc
      121 ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg
      181 cttgaccgac aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga
      241 tgtacgggcc agatatacgc gttgacattg attattgact agttattaat agtaatcaat
      301 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
      361 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
      421 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
      481 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
      541 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
      601 tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca
      661 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
      721 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
      781 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
      841 cagagctctc tggctaacta gagaacccac tgcttactgg cttatcgaaa ttaatacgac
      901 tcactatagg gagacccaag ctggctagcg tttaaactta agcttggtac cgagctcgga
      961 tccactagtc cagtgtggtg gaattctgca gatatccagc acagtggcgg ccgcgccacc
     1021 atgaagacga tcatcgccct gagctacatc ttctgcctgg tattcgccga ctacaaggac
     1081 gatgatgacg ccagcatcga tatggccacg acggtgccag acggctgccg caatggattg
     1141 aaatccaagt attatcgcct ctgtgacaaa gcagaggcat ggggcatagt gctcgaaacg
     1201 gtcgctacag ctggcgtagt aacctccgtg gcatttatgc tcaccctgcc tatcctcgtc
     1261 tgcaaagtgc aggattccaa tagacgcaag atgctgccaa cccagttcct gttcttgctc
     1321 ggcgtgcttg gcatctttgg gctgacgttt gcctttatta tcggcctgga tggctccact
     1381 ggccccacaa ggttctttct gtttgggata ctgttttcta tttgtttctc ctgcctcctg
     1441 gctcatgccg tctcactcac gaagttggtg cgaggaagga aacccctctc actcctcgtg
     1501 atcctcggcc tggctgtcgg cttttccctg gtgcaagatg tgattgcaat cgagtacatc
     1561 gtgttgacaa tgaatcgcac gaacgtcaac gtgttcagcg aactgagcgc tccccggcgg
     1621 aacgaggact tcgtgctgct cctgacttac gtactgtttt tgatggccct gacgttcttg
     1681 atgtcctcct tcaccttttg cgggtccttt actggttgga aaaggcacgg cgcgcatatt
     1741 tatctgacga tgcttctgag tattgctatc tgggtggcct ggatcacttt gctgatgttg
     1801 ccagactttg accgccgctg ggacgacacc atactgagca gtgcactcgc cgccaacggt
     1861 tgggtgtttc tcctcgctta tgtttctccc gagttctggc tgctcacgaa acagcgaaat
     1921 cccatggact acccagtcga ggacgcattc tgcaagcctc agctggtcaa gaagagttat
     1981 ggggtggaga atcgggctta tagccaagag gagatcacac agggtttcga ggagacggga
     2041 gatacacttt atgctccata cagcacacac ttccagctgc agaaccaacc cccccagaag
     2101 gaattttcaa ttccacgcgc ccacgcttgg ccatctccat acaaggacta tgaagtcaag
     2161 aaggaggggt caatcgatac cggtggacgc accccaccca gcctgggtcc ccaagatgag
     2221 tcctgcacca ccgccagctc ctccctggcc aaggacactt catcgaccgg tgagaacctg
     2281 tacttccagc taagattaga taaaagtaaa gtgattaaca gcgcattaga gctgcttaat
     2341 gaggtcggaa tcgaaggttt aacaacccgt aaactcgccc agaagctagg tgtagagcag
     2401 cctacattgt attggcatgt aaaaaataag cgggctttgc tcgacgcctt agccattgag
     2461 atgttagata ggcaccatac tcacttttgc cctttagaag gggaaagctg gcaagatttt
     2521 ttacgtaata acgctaaaag ttttagatgt gctttactaa gtcatcgcga tggagcaaaa
     2581 gtacatttag gtacacggcc tacagaaaaa cagtatgaaa ctctcgaaaa tcaattagcc
     2641 tttttatgcc aacaaggttt ttcactagag aatgcattat atgcactcag cgctgtgggg
     2701 cattttactt taggttgcgt attggaagat caagagcatc aagtcgctaa agaagaaagg
     2761 gaaacaccta ctactgatag tatgccgcca ttattacgac aagctatcga attatttgat
     2821 caccaaggtg cagagccagc cttcttattc ggccttgaat tgatcatatg cggattagaa
     2881 aaacaactta aatgtgaaag tgggtccgcg tacagccgcg cgcgtacgaa aaacaattac
     2941 gggtctacca tcgagggcct gctcgatctc ccggacgacg acgcccccga agaggcgggg
     3001 ctggcggctc cgcgcctgtc ctttctcccc gcgggacaca cgcgcagact gtcgacggcc
     3061 cccccgaccg atgtcagcct gggggacgag ctccacttag acggcgagga cgtggcgatg
     3121 gcgcatgccg acgcgctaga cgatttcgat ctggacatgt tgggggacgg ggattccccg
     3181 ggtccgggat ttacccccca cgactccgcc ccctacggcg ctctggatat ggccgacttc
     3241 gagtttgagc agatgtttac cgatgccctt ggaattgacg agtacggtgg gtagtctaga
     3301 gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     3361 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     3421 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     3481 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat
     3541 gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc
     3601 cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     3661 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     3721 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     3781 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     3841 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     3901 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     3961 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     4021 aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc
     4081 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt
     4141 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     4201 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
     4261 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg
     4321 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc
     4381 cgggagcttg tatatccatt ttcggatctg atcaagagac aggatgagga tcgtttcgca
     4441 tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg
     4501 gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag
     4561 cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc
     4621 aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc
     4681 tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg
     4741 atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc
     4801 ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca
     4861 tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag
     4921 agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg
     4981 gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg
     5041 gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca
     5101 tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc
     5161 tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg
     5221 acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct
     5281 gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt
     5341 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc
     5401 ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     5461 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     5521 tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc
     5581 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
     5641 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     5701 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     5761 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     5821 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     5881 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     5941 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     6001 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     6061 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     6121 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     6181 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     6241 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     6301 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     6361 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     6421 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     6481 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     6541 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     6601 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     6661 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     6721 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     6781 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     6841 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     6901 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     6961 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     7021 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     7081 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     7141 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     7201 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     7261 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     7321 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     7381 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     7441 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     7501 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     7561 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     7621 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     7681 aaataaacaa ataggggttc cgcgca
//
