LOCUS       sequence-347879-        7805 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 7805)
  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 7805)
  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: HTR1B-Tango
FEATURES             Location/Qualifiers
     source          1..7805
                     /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(2129..2743,2744..3130)
                     /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(2196..2215)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(3162..3179)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3168..3392
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3438..3866
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3525..3544)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3735..3756
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3875..3895)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3880..4209
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4060..4195
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4122..4141
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4276..5070
                     /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(4330..4349)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4940..4959
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5244..5365
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5281..5300)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5335..5354
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5414..5430)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5414..5430)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5427..5449)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5438..5454
                     /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(5462..5468,5469..5486,5487..5492))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5507..5528
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5645..5662)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5816..6404)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5896..5915)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6575..7366,7367..7435))
                     /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     7198..7217
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7436..7540)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7615..7634)
                     /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 ggaggagcct ggcgctcaat gcgctccacc tccccctgct gggtctgaga cttgggtgcc
      901 acaggctaat ctgtcaagtg caccaagcca gaactgctcc gcaaaggatt acatatatca
      961 agacagtata agtctgcctt ggaaagtgct tctcgtgatg cttctcgccc tgatcacctt
     1021 ggccacaaca ctctctaacg ccttcgttat agcaaccgtg tatcgaactc ggaagttgca
     1081 tacccccgcc aattatttga ttgcctccct ggccgtgacc gatcttctgg tttcaatcct
     1141 ggtcatgccg atctcaacta tgtacaccgt gactggccga tggacacttg ggcaggtggt
     1201 gtgcgacttc tggttgagct cagacatcac ctgctgtacc gcctctattc tccacctgtg
     1261 cgtcattgct ctggataggt actgggccat caccgacgcc gttgagtata gcgctaagcg
     1321 gacccctaaa agggccgccg tgatgattgc cttggtgtgg gtcttcagca tttcaatctc
     1381 tctcccaccg tttttctgga gacaggctaa agcagaagag gaggtctccg aatgtgtggt
     1441 caacaccgac catatactgt acactgtata ctcaaccgtc ggcgccttct actttcctac
     1501 cctcctcctg atagcactct acggccggat ctatgttgag gcaagaagcc ggatcttgaa
     1561 gcagacgccc aaccgaacag gcaaacgcct cacaagggct caactcatca cagattcacc
     1621 cggctcaaca agctccgtta cctcaatcaa ttctcgggtc cccgatgtcc cgagcgagtc
     1681 cggttctcct gtatacgtga accaggttaa ggtgcgcgta tctgacgcct tgcttgaaaa
     1741 gaaaaaattg atggccgcac gcgaaaggaa agccacaaag actctcggta ttatcctcgg
     1801 agcctttatt gtgtgttggc tgcctttttt cattatcagc ctggtgatgc ctatctgcaa
     1861 agatgcgtgc tggttccacc tcgctatctt cgattttttt acttggctgg ggtacctgaa
     1921 ctctttgatc aatcccatta tttatacaat gtctaacgaa gactttaagc aggcattcca
     1981 taagttgatt cggttcaagt gtacatcaat cgataccggt ggacgcaccc cacccagcct
     2041 gggtccccaa gatgagtcct gcaccaccgc cagctcctcc ctggccaagg acacttcatc
     2101 gaccggtgag aacctgtact tccagctaag attagataaa agtaaagtga ttaacagcgc
     2161 attagagctg cttaatgagg tcggaatcga aggtttaaca acccgtaaac tcgcccagaa
     2221 gctaggtgta gagcagccta cattgtattg gcatgtaaaa aataagcggg ctttgctcga
     2281 cgccttagcc attgagatgt tagataggca ccatactcac ttttgccctt tagaagggga
     2341 aagctggcaa gattttttac gtaataacgc taaaagtttt agatgtgctt tactaagtca
     2401 tcgcgatgga gcaaaagtac atttaggtac acggcctaca gaaaaacagt atgaaactct
     2461 cgaaaatcaa ttagcctttt tatgccaaca aggtttttca ctagagaatg cattatatgc
     2521 actcagcgct gtggggcatt ttactttagg ttgcgtattg gaagatcaag agcatcaagt
     2581 cgctaaagaa gaaagggaaa cacctactac tgatagtatg ccgccattat tacgacaagc
     2641 tatcgaatta tttgatcacc aaggtgcaga gccagccttc ttattcggcc ttgaattgat
     2701 catatgcgga ttagaaaaac aacttaaatg tgaaagtggg tccgcgtaca gccgcgcgcg
     2761 tacgaaaaac aattacgggt ctaccatcga gggcctgctc gatctcccgg acgacgacgc
     2821 ccccgaagag gcggggctgg cggctccgcg cctgtccttt ctccccgcgg gacacacgcg
     2881 cagactgtcg acggcccccc cgaccgatgt cagcctgggg gacgagctcc acttagacgg
     2941 cgaggacgtg gcgatggcgc atgccgacgc gctagacgat ttcgatctgg acatgttggg
     3001 ggacggggat tccccgggtc cgggatttac cccccacgac tccgccccct acggcgctct
     3061 ggatatggcc gacttcgagt ttgagcagat gtttaccgat gcccttggaa ttgacgagta
     3121 cggtgggtag tctagagggc ccgtttaaac ccgctgatca gcctcgactg tgccttctag
     3181 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     3241 tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca
     3301 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag
     3361 caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg
     3421 ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     3481 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
     3541 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     3601 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
     3661 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     3721 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
     3781 ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct
     3841 gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga
     3901 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     3961 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     4021 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     4081 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     4141 gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     4201 ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca agagacagga
     4261 tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg
     4321 gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc
     4381 gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt
     4441 gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt
     4501 ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc
     4561 gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc
     4621 atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac
     4681 caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag
     4741 gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag
     4801 gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat
     4861 atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg
     4921 gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa
     4981 tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc
     5041 ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc
     5101 aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt
     5161 tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca
     5221 tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa
     5281 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     5341 tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct
     5401 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     5461 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     5521 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     5581 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     5641 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     5701 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     5761 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     5821 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     5881 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     5941 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     6001 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     6061 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     6121 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     6181 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     6241 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     6301 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     6361 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     6421 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     6481 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     6541 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     6601 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     6661 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     6721 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     6781 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     6841 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     6901 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     6961 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     7021 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     7081 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     7141 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     7201 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     7261 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     7321 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     7381 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     7441 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     7501 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     7561 cacctgacgt cgacggatcg ggagatctcc cgatccccta tggtgcactc tcagtacaat
     7621 ctgctctgat gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc
     7681 tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat
     7741 gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac
     7801 gcgtt
//
