LOCUS       sequence-344645-        7787 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 7787)
  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 7787)
  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: S1PR4-Tango
FEATURES             Location/Qualifiers
     source          1..7787
                     /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(2111..2725,2726..3112)
                     /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(2178..2197)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(3144..3161)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3150..3374
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3420..3848
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3507..3526)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3717..3738
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3857..3877)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3862..4191
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4042..4177
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4104..4123
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4258..5052
                     /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(4312..4331)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4922..4941
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5226..5347
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5263..5282)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5317..5336
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5396..5412)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5396..5412)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5409..5431)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5420..5436
                     /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(5444..5450,5451..5468,5469..5474))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5489..5510
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5627..5644)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5798..6386)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5878..5897)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6557..7348,7349..7417))
                     /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     7180..7199
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7418..7522)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7597..7616)
                     /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 gaacgctacc ggaacccccg ttgcccctga gagctgtcag caacttgccg ccgggggcca
      901 cagtaggctg atcgtcctgc actacaatca ttccggccga ctggcagggc gcggtggccc
      961 tgaagacggg ggcctcggtg ctctgagggg cctttccgtc gcagctagct gcctggtggt
     1021 tttggaaaac ctccttgtgc tcgccgccat cactagccat atgcgctccc gacgctgggt
     1081 atactactgc ctcgtgaaca ttaccctcag tgacctcctg acaggggccg cctatcttgc
     1141 caatgtgctc ctgtctggcg cccgcacctt ccggcttgct cctgcccaat ggttcctccg
     1201 ggagggcctg ctgttcacag ccctggcggc ttccactttc agcttgctgt tcaccgcagg
     1261 ggagcggttt gctacgatgg tgcgccctgt cgctgagtcc ggcgcaacaa agaccagtcg
     1321 agtttacgga ttcatcggac tctgttggtt gctcgcggca cttctgggca tgctgccact
     1381 tcttgggtgg aactgtttgt gtgcctttga taggtgcagc agcctcctgc cactttacag
     1441 taagagatat attctgtttt gcctcgtgat cttcgccggt gtgttggcca caattatggg
     1501 actgtatggt gctattttta ggcttgtgca ggcctccggt cagaaagcac cccgccccgc
     1561 cgcccggagg aaagcccggc gcctcttgaa aacagtgctg atgattctgc ttgccttcct
     1621 cgtatgctgg ggccccctgt ttgggctgtt gctggcagac gttttcggct caaacctctg
     1681 ggcgcaagag tatctgcgcg ggatggactg gattctggcc ctcgccgtgc tcaacagcgc
     1741 agtgaatcca atcatctact cattcagaag tagggaggtg tgcagggccg tacttagctt
     1801 cctgtgttgc ggctgtctgc gccttggaat gagagggcca ggggattgtc tggccagagc
     1861 cgtagaagca catagtggcg catcaaccac agatagttcc ctcaggccca gggattcttt
     1921 tcgaggcagt cggtccttga gctttagaat gagagaacct ttgtctagta taagctccgt
     1981 tcgctcaatt atcgataccg gtggacgcac cccacccagc ctgggtcccc aagatgagtc
     2041 ctgcaccacc gccagctcct ccctggccaa ggacacttca tcgaccggtg agaacctgta
     2101 cttccagcta agattagata aaagtaaagt gattaacagc gcattagagc tgcttaatga
     2161 ggtcggaatc gaaggtttaa caacccgtaa actcgcccag aagctaggtg tagagcagcc
     2221 tacattgtat tggcatgtaa aaaataagcg ggctttgctc gacgccttag ccattgagat
     2281 gttagatagg caccatactc acttttgccc tttagaaggg gaaagctggc aagatttttt
     2341 acgtaataac gctaaaagtt ttagatgtgc tttactaagt catcgcgatg gagcaaaagt
     2401 acatttaggt acacggccta cagaaaaaca gtatgaaact ctcgaaaatc aattagcctt
     2461 tttatgccaa caaggttttt cactagagaa tgcattatat gcactcagcg ctgtggggca
     2521 ttttacttta ggttgcgtat tggaagatca agagcatcaa gtcgctaaag aagaaaggga
     2581 aacacctact actgatagta tgccgccatt attacgacaa gctatcgaat tatttgatca
     2641 ccaaggtgca gagccagcct tcttattcgg ccttgaattg atcatatgcg gattagaaaa
     2701 acaacttaaa tgtgaaagtg ggtccgcgta cagccgcgcg cgtacgaaaa acaattacgg
     2761 gtctaccatc gagggcctgc tcgatctccc ggacgacgac gcccccgaag aggcggggct
     2821 ggcggctccg cgcctgtcct ttctccccgc gggacacacg cgcagactgt cgacggcccc
     2881 cccgaccgat gtcagcctgg gggacgagct ccacttagac ggcgaggacg tggcgatggc
     2941 gcatgccgac gcgctagacg atttcgatct ggacatgttg ggggacgggg attccccggg
     3001 tccgggattt accccccacg actccgcccc ctacggcgct ctggatatgg ccgacttcga
     3061 gtttgagcag atgtttaccg atgcccttgg aattgacgag tacggtgggt agtctagagg
     3121 gcccgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt
     3181 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
     3241 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
     3301 ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc
     3361 ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca
     3421 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     3481 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     3541 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     3601 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     3661 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     3721 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     3781 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     3841 cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     3901 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     3961 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     4021 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     4081 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag
     4141 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg
     4201 ggagcttgta tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg
     4261 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     4321 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     4381 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag
     4441 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     4501 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     4561 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     4621 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     4681 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     4741 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc
     4801 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     4861 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     4921 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     4981 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     5041 gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc
     5101 catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     5161 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     5221 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     5281 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     5341 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     5401 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     5461 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     5521 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     5581 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     5641 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     5701 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     5761 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     5821 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     5881 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     5941 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     6001 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     6061 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     6121 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     6181 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     6241 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     6301 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     6361 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     6421 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     6481 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     6541 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     6601 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     6661 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     6721 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     6781 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     6841 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     6901 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     6961 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     7021 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     7081 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     7141 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     7201 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     7261 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     7321 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     7381 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     7441 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     7501 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat
     7561 cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg atgccgcata
     7621 gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa
     7681 aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc tgcttagggt
     7741 taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgtt
//
