LOCUS       sequence-494151-        8042 bp    DNA     circular SYN 02-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 8042)
  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 8042)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 2, 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: RXFP3-Tango
FEATURES             Location/Qualifiers
     source          1..8042
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        183..562
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        563..766
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     717..737
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     811..830
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        811..829
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(2548..3162,3163..3549)
                     /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(2615..2634)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(3581..3598)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3587..3811
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3857..4285
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3944..3963)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4154..4175
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4294..4314)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4299..4628
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4479..4614
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4541..4560
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4695..5489
                     /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(4749..4768)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5359..5378
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5663..5784
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5700..5719)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5754..5773
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5833..5849)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5833..5849)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5846..5868)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5857..5873
                     /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(5881..5887,5888..5905,5906..5911))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5926..5947
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6064..6081)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6235..6823)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6315..6334)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6994..7785,7786..7854))
                     /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     7617..7636
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7855..7959)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(join(8034..8042,1..11))
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ctctgatgcc gcatagttaa gccagtatct gctccctgct tgtgtgttgg aggtcgctga
       61 gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc ttgaccgaca attgcatgaa
      121 gaatctgctt agggttaggc gttttgcgct gcttcgcgat gtacgggcca gatatacgcg
      181 ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag
      241 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
      301 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
      361 gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca
      421 tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc
      481 ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt
      541 attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata
      601 gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt
      661 ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca
      721 aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctctct ggctaactag
      781 agaacccact gcttactggc ttatcgaaat taatacgact cactataggg agacccaagc
      841 tggctagcgt ttaaacttaa gcttggtacc gagctcggat ccactagtcc agtgtggtgg
      901 aattctgcag atatccagca cagtggcggc cgcgccacca tgaagacgat catcgccctg
      961 agctacatct tctgcctggt attcgccgac tacaaggacg atgatgacgc cagcatcgat
     1021 atgcagatgg ccgacgccgc cactattgcc actatgaata aggctgcggg tggagacaaa
     1081 ctggcggagc tgttctctct ggtccccgac ctgctggagg ccgctaacac ttcaggcaat
     1141 gcgtcccttc agctcccaga tctttggtgg gaattgggcc tggaactgcc cgacggagcg
     1201 cctcctggcc accctcccgg cagcggcggc gctgaatctg ccgataccga agcccgggtc
     1261 aggatcctga taagcgtcgt ctactgggtt gtgtgcgccc tcggtctggc cggcaatctg
     1321 ctggtgctgt accttatgaa gtcaatgcag gggtggcgca aatcctctat taaccttttc
     1381 gtcaccaacc tcgctctgac cgattttcag tttgtcttga ctctgccctt ctgggcagtt
     1441 gaaaacgccc tggatttcaa atggccattt ggcaaggcca tgtgcaaaat agtgagtatg
     1501 gttacgtcaa tgaacatgta cgcgtcagtg ttctttctga ccgccatgag cgtgacccgg
     1561 tatcatagcg ttgcatccgc cttgaaaagt caccgaaccc gcggtcatgg acggggagat
     1621 tgctgtgggc gaagcctcgg ggacagttgc tgttttagcg ctaaagctct gtgtgtatgg
     1681 atttgggcac tggctgctct cgcatctctc ccttcagcta tctttagtac aactgtgaag
     1741 gtgatgggcg aggagctctg tctcgtgagg tttcctgaca agcttcttgg gcgggataga
     1801 cagttctggc ttgggctgta ccattctcag aaagttctgc tgggctttgt tctgcccctt
     1861 gggatcatta tcctttgtta ccttctgctg gtgcgattta ttgcagaccg acgggctgcc
     1921 gggaccaaag gaggcgccgc agttgcaggc gggcgcccaa caggcgctag cgccagacga
     1981 ctctctaaag ttaccaagag tgttacaatc gtggtactga gtttcttcct ctgctggctt
     2041 cccaaccagg cactgaccac atggagcatc ttgattaaat ttaatgctgt gccgttttca
     2101 caggaatact tcctctgtca ggtctacgca ttccccgtta gtgtctgcct cgcccactca
     2161 aacagctgtc ttaatcctgt gctgtactgc ctggtgagaa gagaattccg aaaggctctg
     2221 aaatcactgc tgtggcggat agcctccccc tctatcacta gcatgcgccc ttttacagcc
     2281 acaaccaaac cagaacacga ggaccagggc ctccaagccc cagctccccc ccacgcagct
     2341 gccgaaccgg accttctgta ctatccaccc ggtgtcgtcg tgtattctgg tggaagatac
     2401 gacttgctcc ccagcagcag cgcatacatc gataccggtg gacgcacccc acccagcctg
     2461 ggtccccaag atgagtcctg caccaccgcc agctcctccc tggccaagga cacttcatcg
     2521 accggtgaga acctgtactt ccagctaaga ttagataaaa gtaaagtgat taacagcgca
     2581 ttagagctgc ttaatgaggt cggaatcgaa ggtttaacaa cccgtaaact cgcccagaag
     2641 ctaggtgtag agcagcctac attgtattgg catgtaaaaa ataagcgggc tttgctcgac
     2701 gccttagcca ttgagatgtt agataggcac catactcact tttgcccttt agaaggggaa
     2761 agctggcaag attttttacg taataacgct aaaagtttta gatgtgcttt actaagtcat
     2821 cgcgatggag caaaagtaca tttaggtaca cggcctacag aaaaacagta tgaaactctc
     2881 gaaaatcaat tagccttttt atgccaacaa ggtttttcac tagagaatgc attatatgca
     2941 ctcagcgctg tggggcattt tactttaggt tgcgtattgg aagatcaaga gcatcaagtc
     3001 gctaaagaag aaagggaaac acctactact gatagtatgc cgccattatt acgacaagct
     3061 atcgaattat ttgatcacca aggtgcagag ccagccttct tattcggcct tgaattgatc
     3121 atatgcggat tagaaaaaca acttaaatgt gaaagtgggt ccgcgtacag ccgcgcgcgt
     3181 acgaaaaaca attacgggtc taccatcgag ggcctgctcg atctcccgga cgacgacgcc
     3241 cccgaagagg cggggctggc ggctccgcgc ctgtcctttc tccccgcggg acacacgcgc
     3301 agactgtcga cggccccccc gaccgatgtc agcctggggg acgagctcca cttagacggc
     3361 gaggacgtgg cgatggcgca tgccgacgcg ctagacgatt tcgatctgga catgttgggg
     3421 gacggggatt ccccgggtcc gggatttacc ccccacgact ccgcccccta cggcgctctg
     3481 gatatggccg acttcgagtt tgagcagatg tttaccgatg cccttggaat tgacgagtac
     3541 ggtgggtagt ctagagggcc cgtttaaacc cgctgatcag cctcgactgt gccttctagt
     3601 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact
     3661 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat
     3721 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc
     3781 aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc
     3841 tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     3901 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
     3961 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     4021 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat
     4081 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     4141 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc
     4201 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
     4261 atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa
     4321 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     4381 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     4441 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     4501 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     4561 ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     4621 tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcaa gagacaggat
     4681 gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg
     4741 tggagaggct attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg
     4801 tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg
     4861 ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc
     4921 cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg
     4981 aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca
     5041 tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc
     5101 aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg
     5161 atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg
     5221 cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata
     5281 tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg
     5341 accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat
     5401 gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct
     5461 tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca
     5521 agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt
     5581 gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat
     5641 gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag
     5701 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     5761 gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt
     5821 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     5881 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     5941 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     6001 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     6061 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     6121 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     6181 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     6241 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     6301 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     6361 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     6421 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     6481 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     6541 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     6601 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     6661 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     6721 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     6781 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     6841 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     6901 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     6961 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     7021 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     7081 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     7141 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     7201 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     7261 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     7321 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     7381 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     7441 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     7501 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     7561 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     7621 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     7681 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     7741 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     7801 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     7861 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     7921 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     7981 acctgacgtc gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc
     8041 tg
//
