LOCUS       sequence-157905-        7772 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 7772)
  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 7772)
  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: OPRK1-Tango
FEATURES             Location/Qualifiers
     source          1..7772
                     /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(2333..2947,2948..3334)
                     /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(2400..2419)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(3366..3383)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3372..3596
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3642..4070
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3729..3748)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3939..3960
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4079..4099)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4084..4413
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4264..4399
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4326..4345
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4480..5274
                     /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(4534..4553)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5144..5163
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5448..5569
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5485..5504)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5539..5558
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5618..5634)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5618..5634)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5631..5653)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5642..5658
                     /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(5666..5672,5673..5690,5691..5696))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5711..5732
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5849..5866)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6020..6605)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6100..6119)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6776..7567,7568..7636))
                     /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     7399..7418
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7637..7741)
                     /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 atatggacag
     1081 ccccattcaa atcttccggg gcgagcccgg accaacctgc gctccaagcg cttgcctgcc
     1141 accgaattct agtgcatggt tcccagggtg ggcagagccc gatagcaatg gctcagccgg
     1201 gtccgaggat gcgcagcttg aacccgctca catctcaccc gctatccctg tcattattac
     1261 ggcagtctac tcagttgttt tcgttgtggg cctggttgga aatagcctcg tgatgtttgt
     1321 gattatcagg tatactaaaa tgaagaccgc cacgaatatc tacatcttta acctggctct
     1381 ggctgacgcc ttggtgacca ccactatgcc atttcagtcc accgtctatc ttatgaactc
     1441 ttggccattc ggcgacgttc tgtgcaagat tgtcatttcc attgactatt ataatatgtt
     1501 tacatcaatc tttacgctta caatgatgag cgtggaccga tacattgctg tttgccatcc
     1561 ggtgaaagca ctggattttc gcaccccctt gaaggccaag attatcaata tctgtatttg
     1621 gctcctgtcc tcatccgtcg gaatttccgc cattgtcttg ggtggtacaa aggtgaggga
     1681 ggacgtggac gttatagagt gcagcctgca gttccctgac gatgactaca gctggtggga
     1741 cctttttatg aagatctgtg tcttcatttt tgctttcgtc atccccgtgc tgattatcat
     1801 agtgtgctat acactgatga tattgcggct gaaatctgtg cgcctgctga gcggcagccg
     1861 cgaaaaggat cggaatctgc gccgaatcac acgcctcgtg ctggttgtag tcgctgtctt
     1921 cgtggtatgc tggaccccaa tccacatttt catcctggtg gaagctctgg gatccacctc
     1981 tcactccact gccgcactta gctcatatta cttctgcatt gccttgggct acactaattc
     2041 ctcactgaat cccattctgt acgcatttct tgacgagaac ttcaaacggt gctttagaga
     2101 cttttgcttc cccctcaaga tgcgcatgga gcggcagtca accagccgag tccggaatac
     2161 ggtccaggat cccgcctatc tgcgggacat cgacggcatg aacaagcctg tgatcgatac
     2221 cggtggacgc accccaccca gcctgggtcc ccaagatgag tcctgcacca ccgccagctc
     2281 ctccctggcc aaggacactt catcgaccgg tgagaacctg tacttccagc taagattaga
     2341 taaaagtaaa gtgattaaca gcgcattaga gctgcttaat gaggtcggaa tcgaaggttt
     2401 aacaacccgt aaactcgccc agaagctagg tgtagagcag cctacattgt attggcatgt
     2461 aaaaaataag cgggctttgc tcgacgcctt agccattgag atgttagata ggcaccatac
     2521 tcacttttgc cctttagaag gggaaagctg gcaagatttt ttacgtaata acgctaaaag
     2581 ttttagatgt gctttactaa gtcatcgcga tggagcaaaa gtacatttag gtacacggcc
     2641 tacagaaaaa cagtatgaaa ctctcgaaaa tcaattagcc tttttatgcc aacaaggttt
     2701 ttcactagag aatgcattat atgcactcag cgctgtgggg cattttactt taggttgcgt
     2761 attggaagat caagagcatc aagtcgctaa agaagaaagg gaaacaccta ctactgatag
     2821 tatgccgcca ttattacgac aagctatcga attatttgat caccaaggtg cagagccagc
     2881 cttcttattc ggccttgaat tgatcatatg cggattagaa aaacaactta aatgtgaaag
     2941 tgggtccgcg tacagccgcg cgcgtacgaa aaacaattac gggtctacca tcgagggcct
     3001 gctcgatctc ccggacgacg acgcccccga agaggcgggg ctggcggctc cgcgcctgtc
     3061 ctttctcccc gcgggacaca cgcgcagact gtcgacggcc cccccgaccg atgtcagcct
     3121 gggggacgag ctccacttag acggcgagga cgtggcgatg gcgcatgccg acgcgctaga
     3181 cgatttcgat ctggacatgt tgggggacgg ggattccccg ggtccgggat ttacccccca
     3241 cgactccgcc ccctacggcg ctctggatat ggccgacttc gagtttgagc agatgtttac
     3301 cgatgccctt ggaattgacg agtacggtgg gtagtctaga gggcccgttt aaacccgctg
     3361 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     3421 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     3481 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     3541 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
     3601 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     3661 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     3721 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     3781 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     3841 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     3901 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     3961 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     4021 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
     4081 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     4141 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     4201 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     4261 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     4321 ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga
     4381 ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt
     4441 ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     4501 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     4561 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     4621 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta
     4681 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     4741 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     4801 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     4861 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     4921 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     4981 gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc
     5041 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     5101 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     5161 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     5221 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     5281 ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     5341 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     5401 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg
     5461 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     5521 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta
     5581 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     5641 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     5701 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     5761 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     5821 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     5881 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     5941 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     6001 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     6061 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     6121 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     6181 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     6241 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     6301 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     6361 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     6421 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     6481 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     6541 ccaccgctgg tagcggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     6601 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     6661 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     6721 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     6781 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     6841 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     6901 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     6961 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     7021 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     7081 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     7141 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     7201 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     7261 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     7321 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     7381 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     7441 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     7501 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     7561 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     7621 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     7681 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     7741 gcacatttcc ccgaaaagtg ccacctgacg tc
//
