LOCUS       sequence-417865-        8030 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 8030)
  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 8030)
  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: ADRA2A-Tango
FEATURES             Location/Qualifiers
     source          1..8030
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(2354..2968,2969..3355)
                     /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(2421..2440)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(3387..3404)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3393..3617
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3663..4091
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3750..3769)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3960..3981
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4100..4120)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4105..4434
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4285..4420
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4347..4366
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4501..5295
                     /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(4555..4574)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5165..5184
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5469..5590
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5506..5525)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5560..5579
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5639..5655)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5639..5655)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5652..5674)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5663..5679
                     /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(5687..5693,5694..5711,5712..5717))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5732..5753
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5870..5887)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6041..6629)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6121..6140)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6800..7591,7592..7660))
                     /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     7423..7442
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7661..7765)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7840..7859)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg
      661 gctagcgttt aaacttaagc ttggtaccga gctcggatcc actagtccag tgtggtggaa
      721 ttctgcagat atccagcaca gtggcggccg cgccaccatg aagacgatca tcgccctgag
      781 ctacatcttc tgcctggtat tcgccgacta caaggacgat gatgacgcca gcatcgatat
      841 gttccggcag gagcagccac tcgccgaggg ttcctttgcc cccatgggaa gtcttcagcc
      901 tgacgccggc aacgctagct ggaatggcac cgaagcgccc ggtggagggg ctcgggctac
      961 cccatattct ctccaggtca cactgaccct ggtttgtctg gccggtctcc tgatgctgct
     1021 gacggtcttt ggcaatgttt tggtgataat tgcggtgttc acctccagag cactcaaggc
     1081 tccccagaac ctgttcttgg tcagcctggc gtctgccgat attctggtcg ccaccctcgt
     1141 aattccgttt tctcttgcca atgaggttat gggttactgg tattttggta aggcatggtg
     1201 cgagatatac cttgctctcg acgttctctt ctgcaccagc agtattgtgc atctgtgcgc
     1261 aattagtctg gacagatatt ggtccatcac gcaggcaata gaatataact tgaaaaggac
     1321 tcctcggcgc attaaggcta tcatcatcac tgtctgggtc attagcgctg tgataagctt
     1381 ccctccactt atttccattg aaaaaaaagg aggaggcgga gggccgcagc ctgcggagcc
     1441 ccgctgtgag atcaacgatc agaaatggta tgtcatctcc tcctgcattg ggtccttctt
     1501 cgccccatgt ctgatcatga tcctcgtata tgttaggatt tatcagatag ctaagcgaag
     1561 gacccgggtc cccccatcca ggagaggtcc agacgcagtg gccgctcccc ccggaggcac
     1621 agaaaggaga cctaacggtc tgggaccaga gcgctccgca ggacctggag gtgcagaagc
     1681 cgagcccttg ccgacacaac tgaacggagc tcccggcgaa ccagccccag caggtcctcg
     1741 cgatactgac gcactggacc tcgaagagtc ctctagcagc gatcatgcag agcgccctcc
     1801 agggcctagg agacctgaaa gaggacccag aggtaaaggc aaggcccggg caagtcaagt
     1861 gaaaccggga gactctctgc ctcggagggg tccaggtgct actggtatag ggactcctgc
     1921 cgccggtccc ggagaagagc gggtcggcgc cgctaaggca agtcgctggc ggggcagaca
     1981 aaaccgggaa aaaaggttta cattcgtgct cgcggtagtg attggcgtct tcgtcgtgtg
     2041 ctggtttcca ttctttttta cctataccct gacagccgtc ggctgttctg ttccacggac
     2101 actgttcaag ttctttttct ggtttggcta ctgtaactca agcctgaacc cggtaatcta
     2161 tacaatattt aaccacgatt ttcggagggc ttttaaaaag atcctgtgca gaggcgatag
     2221 gaaacgaatc gttatcgata ccggtggacg caccccaccc agcctgggtc cccaagatga
     2281 gtcctgcacc accgccagct cctccctggc caaggacact tcatcgaccg gtgagaacct
     2341 gtacttccag ctaagattag ataaaagtaa agtgattaac agcgcattag agctgcttaa
     2401 tgaggtcgga atcgaaggtt taacaacccg taaactcgcc cagaagctag gtgtagagca
     2461 gcctacattg tattggcatg taaaaaataa gcgggctttg ctcgacgcct tagccattga
     2521 gatgttagat aggcaccata ctcacttttg ccctttagaa ggggaaagct ggcaagattt
     2581 tttacgtaat aacgctaaaa gttttagatg tgctttacta agtcatcgcg atggagcaaa
     2641 agtacattta ggtacacggc ctacagaaaa acagtatgaa actctcgaaa atcaattagc
     2701 ctttttatgc caacaaggtt tttcactaga gaatgcatta tatgcactca gcgctgtggg
     2761 gcattttact ttaggttgcg tattggaaga tcaagagcat caagtcgcta aagaagaaag
     2821 ggaaacacct actactgata gtatgccgcc attattacga caagctatcg aattatttga
     2881 tcaccaaggt gcagagccag ccttcttatt cggccttgaa ttgatcatat gcggattaga
     2941 aaaacaactt aaatgtgaaa gtgggtccgc gtacagccgc gcgcgtacga aaaacaatta
     3001 cgggtctacc atcgagggcc tgctcgatct cccggacgac gacgcccccg aagaggcggg
     3061 gctggcggct ccgcgcctgt cctttctccc cgcgggacac acgcgcagac tgtcgacggc
     3121 ccccccgacc gatgtcagcc tgggggacga gctccactta gacggcgagg acgtggcgat
     3181 ggcgcatgcc gacgcgctag acgatttcga tctggacatg ttgggggacg gggattcccc
     3241 gggtccggga tttacccccc acgactccgc cccctacggc gctctggata tggccgactt
     3301 cgagtttgag cagatgttta ccgatgccct tggaattgac gagtacggtg ggtagtctag
     3361 agggcccgtt taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
     3421 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     3481 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     3541 tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga
     3601 tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc
     3661 ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     3721 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     3781 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     3841 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
     3901 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     3961 tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt
     4021 ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
     4081 taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
     4141 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
     4201 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     4261 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     4321 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct
     4381 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc
     4441 ccgggagctt gtatatccat tttcggatct gatcaagaga caggatgagg atcgtttcgc
     4501 atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc
     4561 ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca
     4621 gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg
     4681 caggacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg
     4741 ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag
     4801 gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg
     4861 cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc
     4921 atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa
     4981 gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac
     5041 ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat
     5101 ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac
     5161 atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc
     5221 ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt
     5281 gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc
     5341 tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg
     5401 ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg
     5461 cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     5521 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     5581 atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt
     5641 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     5701 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     5761 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     5821 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     5881 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     5941 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     6001 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     6061 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     6121 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     6181 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     6241 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     6301 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     6361 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     6421 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     6481 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     6541 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     6601 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     6661 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     6721 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     6781 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     6841 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     6901 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     6961 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     7021 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     7081 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     7141 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     7201 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     7261 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     7321 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     7381 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     7441 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     7501 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     7561 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     7621 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     7681 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     7741 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtcgacg
     7801 gatcgggaga tctcccgatc ccctatggtg cactctcagt acaatctgct ctgatgccgc
     7861 atagttaagc cagtatctgc tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag
     7921 caaaatttaa gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga atctgcttag
     7981 ggttaggcgt tttgcgctgc ttcgcgatgt acgggccaga tatacgcgtt
//
