LOCUS       sequence-344626-        7694 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 7694)
  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 7694)
  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: S1PR2-Tango
FEATURES             Location/Qualifiers
     source          1..7694
                     /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(2018..2632,2633..3019)
                     /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(2085..2104)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(3051..3068)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3057..3281
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3327..3755
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3414..3433)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3624..3645
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3764..3784)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3769..4098
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3949..4084
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4011..4030
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4165..4959
                     /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(4219..4238)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4829..4848
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5133..5254
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5170..5189)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5224..5243
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5303..5319)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5303..5319)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5316..5338)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5327..5343
                     /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(5351..5357,5358..5375,5376..5381))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5396..5417
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5534..5551)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5705..6293)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5785..5804)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6464..7255,7256..7324))
                     /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     7087..7106
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7325..7429)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7504..7523)
                     /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 gggctcactt tacagtgaat acctgaatcc caacaaggtc caagaacact acaattatac
      901 caaggaaaca ctggagacac aggaaacgac ttcccgacag gtggcctccg cattcattgt
      961 gattctctgc tgtgcaatcg tggtggagaa tctgcttgtg ctgatcgcag tagctaggaa
     1021 tagcaaattt catagcgcta tgtatctctt ccttggaaat ctggccgctt ctgaccttct
     1081 tgcgggtgtt gccttcgtgg caaatacact gttgtccggg tccgttacat tgcggctcac
     1141 ccccgtgcag tggtttgccc gcgagggctc cgcctttatc actctgtccg cttccgtctt
     1201 cagcctcctg gcgatcgcca ttgagagaca tgtggcaatc gcgaaagtga aactgtacgg
     1261 gagtgacaag agctgcagaa tgctgctgct gatcggcgca agctggttga tcagtctggt
     1321 tttgggcggc ctgcccatcc tcgggtggaa ctgtctcggt catctcgaag catgtagcac
     1381 tgtgttgcca ctgtacgcaa aacattacgt cctgtgcgtt gttactatct ttagcattat
     1441 tctgcttgca atcgtggcac tgtacgtaag gatatactgt gtggtcagaa gttctcacgc
     1501 tgatatggca gcacctcaga ccctcgcact cctgaaaaca gtcaccattg tgctgggagt
     1561 gttcatagtg tgttggctgc ccgcgttcag catattgctc ttggactacg cctgtcccgt
     1621 gcattcctgc cccatcttgt ataaggccca ttactttttt gctgtgtcca cccttaactc
     1681 cctgctcaat ccagttattt acacatggcg atctcgcgat ctgcgacgag aagtgcttcg
     1741 gcccctccag tgttggaggc ctggggttgg agttcagggc agaagacgcg gaggaactcc
     1801 aggtcaccac ctgttgcccc tgaggtctag tagtagcttg gaacgcggta tgcatatgcc
     1861 cacttctccc accttccttg agggcaacac cgtcgtgatc gataccggtg gacgcacccc
     1921 acccagcctg ggtccccaag atgagtcctg caccaccgcc agctcctccc tggccaagga
     1981 cacttcatcg accggtgaga acctgtactt ccagctaaga ttagataaaa gtaaagtgat
     2041 taacagcgca ttagagctgc ttaatgaggt cggaatcgaa ggtttaacaa cccgtaaact
     2101 cgcccagaag ctaggtgtag agcagcctac attgtattgg catgtaaaaa ataagcgggc
     2161 tttgctcgac gccttagcca ttgagatgtt agataggcac catactcact tttgcccttt
     2221 agaaggggaa agctggcaag attttttacg taataacgct aaaagtttta gatgtgcttt
     2281 actaagtcat cgcgatggag caaaagtaca tttaggtaca cggcctacag aaaaacagta
     2341 tgaaactctc gaaaatcaat tagccttttt atgccaacaa ggtttttcac tagagaatgc
     2401 attatatgca ctcagcgctg tggggcattt tactttaggt tgcgtattgg aagatcaaga
     2461 gcatcaagtc gctaaagaag aaagggaaac acctactact gatagtatgc cgccattatt
     2521 acgacaagct atcgaattat ttgatcacca aggtgcagag ccagccttct tattcggcct
     2581 tgaattgatc atatgcggat tagaaaaaca acttaaatgt gaaagtgggt ccgcgtacag
     2641 ccgcgcgcgt acgaaaaaca attacgggtc taccatcgag ggcctgctcg atctcccgga
     2701 cgacgacgcc cccgaagagg cggggctggc ggctccgcgc ctgtcctttc tccccgcggg
     2761 acacacgcgc agactgtcga cggccccccc gaccgatgtc agcctggggg acgagctcca
     2821 cttagacggc gaggacgtgg cgatggcgca tgccgacgcg ctagacgatt tcgatctgga
     2881 catgttgggg gacggggatt ccccgggtcc gggatttacc ccccacgact ccgcccccta
     2941 cggcgctctg gatatggccg acttcgagtt tgagcagatg tttaccgatg cccttggaat
     3001 tgacgagtac ggtgggtagt ctagagggcc cgtttaaacc cgctgatcag cctcgactgt
     3061 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     3121 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     3181 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     3241 agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac
     3301 cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg
     3361 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     3421 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     3481 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     3541 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     3601 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     3661 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     3721 aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta
     3781 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     3841 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     3901 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     3961 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     4021 gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     4081 ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcaa
     4141 gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg
     4201 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
     4261 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
     4321 ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg
     4381 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
     4441 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
     4501 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
     4561 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
     4621 gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc
     4681 aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
     4741 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
     4801 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt
     4861 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
     4921 cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa
     4981 tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct
     5041 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     5101 gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt
     5161 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     5221 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta
     5281 gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     5341 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     5401 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     5461 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     5521 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     5581 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     5641 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     5701 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     5761 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     5821 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     5881 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     5941 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     6001 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     6061 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     6121 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     6181 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     6241 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     6301 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     6361 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     6421 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     6481 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     6541 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     6601 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     6661 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     6721 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     6781 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     6841 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     6901 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     6961 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     7021 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     7081 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     7141 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     7201 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     7261 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     7321 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     7381 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     7441 gaaaagtgcc acctgacgtc gacggatcgg gagatctccc gatcccctat ggtgcactct
     7501 cagtacaatc tgctctgatg ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt
     7561 ggaggtcgct gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag gcttgaccga
     7621 caattgcatg aagaatctgc ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc
     7681 cagatatacg cgtt
//
