LOCUS       sequence-456564-        7955 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 7955)
  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 7955)
  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: HTR6-Tango
FEATURES             Location/Qualifiers
     source          1..7955
                     /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(2513..3127,3128..3514)
                     /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(2580..2599)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(3546..3563)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3552..3776
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3822..4250
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3909..3928)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4119..4140
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4259..4279)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4264..4593
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4444..4579
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4506..4525
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4660..5454
                     /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(4714..4733)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5324..5343
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5628..5749
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5665..5684)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5719..5738
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5798..5814)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5798..5814)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5811..5833)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5822..5838
                     /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(5846..5852,5853..5870,5871..5876))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5891..5912
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6029..6046)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6200..6788)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6280..6299)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6959..7750,7751..7819))
                     /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     7582..7601
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7820..7924)
                     /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 atatggtgcc
     1081 cgagccgggc ccaactgcta acagcacccc agcttggggg gctggtcctc caagcgcccc
     1141 aggtggcagt ggttgggttg ctgcagcatt gtgcgtggtt atcgccctga cagctgccgc
     1201 aaacagcctg ctcattgctc ttatttgtac ccaacccgca ctccggaata cgtctaattt
     1261 tttcctggtc tcactgttca cgagcgacct tatggtcggg ctcgtagtca tgcctcctgc
     1321 tatgttgaac gccctgtatg ggagatgggt cctggcaagg ggtctttgcc ttctgtggac
     1381 cgcatttgac gtcatgtgct gtagtgccag catcttgaac ctctgtttga ttagtctgga
     1441 tagatatctg ctgattctga gcccacttag atacaagctg cggatgactc cgctccgggc
     1501 cctcgctttg gtgctgggcg cctggtctct ggcagcactg gcttcattct tgcctttgct
     1561 gctgggctgg cacgaattgg ggcacgcaag gccgccagtt ccaggccagt gtaggctgct
     1621 tgcctccctg cccttcgtgt tggtggcttc cggcctcaca ttctttctgc ctagtggtgc
     1681 catctgcttc acttactgcc ggatacttct ggcagctaga aaacaagccg ttcaggttgc
     1741 ctccctcacc accggcatgg caagccaggc ctctgagacc ctgcaggtgc cgaggactcc
     1801 taggcctggc gtagagtctg ccgactcacg gcggctggcc actaaacact caagaaaggc
     1861 gctgaaagct tcattgacct tgggcatctt gctgggcatg ttcttcgtaa catggctccc
     1921 attttttgtg gccaatattg tccaggcggt gtgcgactgc atctcaccag gcctgttcga
     1981 tgtgctcact tggctggggt attgcaattc cactatgaac cccatcatat atcccctgtt
     2041 catgagagat ttcaagcgag cactcggaag gtttctgcca tgcccacggt gcccacgcga
     2101 aaggcaagca agtctggcct ccccctcact ccggacctcc cattcaggac ccagaccagg
     2161 actttcactc cagcaagttc tgcccctgcc acttcctcca gattcagact ctgactccga
     2221 tgcgggttca ggtggctcta gcggactccg gcttacggcc cagcttttgc ttcccgggga
     2281 agcaactcag gacccaccat tgcctactcg ggctgccgcc gccgtcaatt tcttcaacat
     2341 tgatcctgca gagccagaac tcagaccgca tcctctgggt ataccaacca atatcgatac
     2401 cggtggacgc accccaccca gcctgggtcc ccaagatgag tcctgcacca ccgccagctc
     2461 ctccctggcc aaggacactt catcgaccgg tgagaacctg tacttccagc taagattaga
     2521 taaaagtaaa gtgattaaca gcgcattaga gctgcttaat gaggtcggaa tcgaaggttt
     2581 aacaacccgt aaactcgccc agaagctagg tgtagagcag cctacattgt attggcatgt
     2641 aaaaaataag cgggctttgc tcgacgcctt agccattgag atgttagata ggcaccatac
     2701 tcacttttgc cctttagaag gggaaagctg gcaagatttt ttacgtaata acgctaaaag
     2761 ttttagatgt gctttactaa gtcatcgcga tggagcaaaa gtacatttag gtacacggcc
     2821 tacagaaaaa cagtatgaaa ctctcgaaaa tcaattagcc tttttatgcc aacaaggttt
     2881 ttcactagag aatgcattat atgcactcag cgctgtgggg cattttactt taggttgcgt
     2941 attggaagat caagagcatc aagtcgctaa agaagaaagg gaaacaccta ctactgatag
     3001 tatgccgcca ttattacgac aagctatcga attatttgat caccaaggtg cagagccagc
     3061 cttcttattc ggccttgaat tgatcatatg cggattagaa aaacaactta aatgtgaaag
     3121 tgggtccgcg tacagccgcg cgcgtacgaa aaacaattac gggtctacca tcgagggcct
     3181 gctcgatctc ccggacgacg acgcccccga agaggcgggg ctggcggctc cgcgcctgtc
     3241 ctttctcccc gcgggacaca cgcgcagact gtcgacggcc cccccgaccg atgtcagcct
     3301 gggggacgag ctccacttag acggcgagga cgtggcgatg gcgcatgccg acgcgctaga
     3361 cgatttcgat ctggacatgt tgggggacgg ggattccccg ggtccgggat ttacccccca
     3421 cgactccgcc ccctacggcg ctctggatat ggccgacttc gagtttgagc agatgtttac
     3481 cgatgccctt ggaattgacg agtacggtgg gtagtctaga gggcccgttt aaacccgctg
     3541 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     3601 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     3661 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     3721 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
     3781 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     3841 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     3901 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     3961 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     4021 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     4081 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     4141 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     4201 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
     4261 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     4321 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     4381 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     4441 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     4501 ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga
     4561 ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt
     4621 ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     4681 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     4741 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     4801 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta
     4861 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     4921 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     4981 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     5041 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     5101 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     5161 gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc
     5221 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     5281 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     5341 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     5401 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     5461 ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     5521 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     5581 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg
     5641 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     5701 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta
     5761 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     5821 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     5881 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     5941 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     6001 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     6061 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     6121 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     6181 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     6241 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     6301 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     6361 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     6421 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     6481 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     6541 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     6601 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     6661 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     6721 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     6781 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     6841 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     6901 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     6961 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     7021 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     7081 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     7141 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     7201 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     7261 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     7321 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     7381 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     7441 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     7501 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     7561 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     7621 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     7681 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     7741 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     7801 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     7861 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     7921 cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
