LOCUS       sequence-163593-        7715 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 7715)
  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 7715)
  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: AGTR1-Tango
FEATURES             Location/Qualifiers
     source          1..7715
                     /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(2270..2884,2885..3271)
                     /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(2337..2356)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(3309..3326)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3315..3539
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3585..4013
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3672..3691)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3882..3903
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4022..4042)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4027..4356
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4207..4342
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4269..4288
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4423..5217
                     /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(4477..4496)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5087..5106
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5391..5512
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5428..5447)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5482..5501
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5561..5577)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5561..5577)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5574..5596)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5585..5601
                     /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(5609..5615,5616..5633,5634..5639))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5654..5675
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5792..5809)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5963..6548)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6043..6062)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6719..7510,7511..7579))
                     /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     7342..7361
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7580..7684)
                     /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 atatgatcct
     1081 caactcctca actgaagacg gtataaaaag aatccaggat gattgcccaa aagcagggcg
     1141 gcataactac atattcgtga tgatccccac tctttatagt attatctttg tggtgggcat
     1201 cttcgggaat tcactcgttg tgatcgtgat ttacttttat atgaagctta agacggtggc
     1261 ctccgttttc ctgctgaatc tcgctttggc cgacctgtgc tttcttctca ccttgccctt
     1321 gtgggccgtg tacaccgcga tggagtaccg atggcctttc ggtaactacc tgtgtaagat
     1381 cgctagcgcc agcgtatctt ttaatcttta tgcctctgtt ttcctcctca catgcctgtc
     1441 tattgatcgg tatctggcca tcgtacatcc gatgaaatca agacttagga ggacaatgct
     1501 ggttgctaag gtaacctgta ttattatttg gctgctggca ggtctggctt ccctgcctgc
     1561 aattattcat cgcaacgtct ttttcatcga gaacaccaac attaccgtct gcgcctttca
     1621 ctacgagagt caaaattcca cgctcccaat aggactgggc ctgacaaaaa acattctcgg
     1681 ttttctcttt ccattcctca tcatcttgac ctcatatacc ttgatctgga aggcactgaa
     1741 aaaggcctat gagatccaga aaaacaaacc cagaaacgac gatattttta aaatcatcat
     1801 ggctattgtg ctgttttttt tcttcagttg gatcccacac cagatcttta ccttccttga
     1861 cgtcttgatt caactgggca tcatcagaga ttgccgcatt gccgacatcg tggacacggc
     1921 catgcccatt acgatttgca tcgcctactt taataactgc cttaatcccc tgttctacgg
     1981 gttcctcggc aagaaattca agcggtactt tcttcagctg ctcaaataca tcccgcccaa
     2041 ggcaaaaagc cactcaaatc tgtcaaccaa gatgagtaca ctgagttaca gaccatccga
     2101 caacgtgagc agttctacca agaaacccgc accatgtttc gaggtcgaaa tcgataccgg
     2161 tggacgcacc ccacccagcc tgggtcccca agatgagtcc tgcaccaccg ccagctcctc
     2221 cctggccaag gacacttcat cgaccggtga gaacctgtac ttccagctaa gattagataa
     2281 aagtaaagtg attaacagcg cattagagct gcttaatgag gtcggaatcg aaggtttaac
     2341 aacccgtaaa ctcgcccaga agctaggtgt agagcagcct acattgtatt ggcatgtaaa
     2401 aaataagcgg gctttgctcg acgccttagc cattgagatg ttagataggc accatactca
     2461 cttttgccct ttagaagggg aaagctggca agatttttta cgtaataacg ctaaaagttt
     2521 tagatgtgct ttactaagtc atcgcgatgg agcaaaagta catttaggta cacggcctac
     2581 agaaaaacag tatgaaactc tcgaaaatca attagccttt ttatgccaac aaggtttttc
     2641 actagagaat gcattatatg cactcagcgc tgtggggcat tttactttag gttgcgtatt
     2701 ggaagatcaa gagcatcaag tcgctaaaga agaaagggaa acacctacta ctgatagtat
     2761 gccgccatta ttacgacaag ctatcgaatt atttgatcac caaggtgcag agccagcctt
     2821 cttattcggc cttgaattga tcatatgcgg attagaaaaa caacttaaat gtgaaagtgg
     2881 gtccgcgtac agccgcgcgc gtacgaaaaa caattacggg tctaccatcg agggcctgct
     2941 cgatctcccg gacgacgacg cccccgaaga ggcggggctg gcggctccgc gcctgtcctt
     3001 tctccccgcg ggacacacgc gcagactgtc gacggccccc ccgaccgatg tcagcctggg
     3061 ggacgagctc cacttagacg gcgaggacgt ggcgatggcg catgccgacg cgctagacga
     3121 tttcgatctg gacatgttgg gggacgggga ttccccgggt ccgggattta ccccccacga
     3181 ctccgccccc tacggcgctc tggatatggc cgacttcgag tttgagcaga tgtttaccga
     3241 tgcccttgga attgacgagt acggtgggta gctcgagtct agagggcccg tttaaacccg
     3301 ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     3361 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     3421 tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag
     3481 caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc
     3541 ttctgaggcg gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg
     3601 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     3661 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     3721 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     3781 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     3841 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     3901 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     3961 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attaattctg
     4021 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     4081 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     4141 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     4201 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     4261 atttttttta tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag
     4321 tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc
     4381 attttcggat ctgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga
     4441 ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa
     4501 cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt
     4561 ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg
     4621 ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa
     4681 gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac
     4741 cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt
     4801 gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact
     4861 cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg
     4921 ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg
     4981 acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc
     5041 atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt
     5101 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc
     5161 gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg
     5221 ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg
     5281 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     5341 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta
     5401 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     5461 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     5521 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     5581 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     5641 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     5701 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     5761 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     5821 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     5881 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     5941 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     6001 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     6061 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     6121 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     6181 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     6241 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     6301 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     6361 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     6421 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     6481 aaaccaccgc tggtagcggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     6541 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     6601 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     6661 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     6721 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     6781 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     6841 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     6901 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     6961 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     7021 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     7081 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     7141 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     7201 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     7261 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     7321 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     7381 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     7441 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     7501 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     7561 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     7621 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     7681 cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
