LOCUS       sequence-314158-        7700 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 7700)
  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 7700)
  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: CCR3-Tango
FEATURES             Location/Qualifiers
     source          1..7700
                     /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(2024..2638,2639..3025)
                     /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(2091..2110)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(3057..3074)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3063..3287
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3333..3761
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3420..3439)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3630..3651
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3770..3790)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3775..4104
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3955..4090
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4017..4036
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4171..4965
                     /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(4225..4244)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4835..4854
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5139..5260
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5176..5195)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5230..5249
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5309..5325)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5309..5325)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5322..5344)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5333..5349
                     /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(5357..5363,5364..5381,5382..5387))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5402..5423
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5540..5557)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5711..6299)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5791..5810)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6470..7261,7262..7330))
                     /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     7093..7112
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7331..7435)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7510..7529)
                     /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 gacaacatct ctggatactg ttgagacttt tggaacgacc tcttactacg atgatgtcgg
      901 tctgttgtgc gagaaggcag acacgcgggc cctcatggca cagttcgtgc cccccctcta
      961 tagccttgtc tttacggtgg ggctgctcgg aaatgtggtg gtggttatga tcttgatcaa
     1021 gtacagacgc cttcggatta tgacgaacat ttatctgctg aacctggcaa tcagcgatct
     1081 tctgttcctc gtgaccctcc ccttttggat acattacgtg cgggggcata actgggtgtt
     1141 tgggcacggt atgtgcaagt tgcttagtgg gttctaccac accgggctct acagcgagat
     1201 attcttcata atcctgctga ccattgatcg gtatcttgcc atcgtgcacg ctgtctttgc
     1261 tcttcgcgct cgcactgtaa cctttggcgt gataacaagt atcgtgactt ggggcctcgc
     1321 tgtgctggct gccctgccag agtttatttt ctatgagacc gaagaacttt tcgaggaaac
     1381 tttgtgctca gctctctacc ccgaagatac agtgtatagt tggcgacatt tccacactct
     1441 ccggatgaca atattctgcc ttgtcctgcc gcttctggtg atggctatct gttataccgg
     1501 aatcataaag accctgctca gatgcccatc taaaaagaag tacaaggcca tccggctcat
     1561 ctttgtgatc atggcagttt tctttatttt ttggacgccc tacaatgttg ctattttgct
     1621 gtccagctac caatccatcc tgtttggcaa cgattgcgaa cggagcaagc acctcgacct
     1681 ggtgatgctt gttaccgagg tcattgccta ttctcactgc tgcatgaatc ccgtaatcta
     1741 tgctttcgta ggcgagagat tcagaaagta cttgcggcat ttcttccaca gacatctgct
     1801 gatgcatctg ggtcgctata tcccttttct gcctagtgaa aagctggaac gcacctcatc
     1861 agtcagcccg agcactgcag agcccgagct gagcatcgtc tttatcgata ccggtggacg
     1921 caccccaccc agcctgggtc cccaagatga gtcctgcacc accgccagct cctccctggc
     1981 caaggacact tcatcgaccg gtgagaacct gtacttccag ctaagattag ataaaagtaa
     2041 agtgattaac agcgcattag agctgcttaa tgaggtcgga atcgaaggtt taacaacccg
     2101 taaactcgcc cagaagctag gtgtagagca gcctacattg tattggcatg taaaaaataa
     2161 gcgggctttg ctcgacgcct tagccattga gatgttagat aggcaccata ctcacttttg
     2221 ccctttagaa ggggaaagct ggcaagattt tttacgtaat aacgctaaaa gttttagatg
     2281 tgctttacta agtcatcgcg atggagcaaa agtacattta ggtacacggc ctacagaaaa
     2341 acagtatgaa actctcgaaa atcaattagc ctttttatgc caacaaggtt tttcactaga
     2401 gaatgcatta tatgcactca gcgctgtggg gcattttact ttaggttgcg tattggaaga
     2461 tcaagagcat caagtcgcta aagaagaaag ggaaacacct actactgata gtatgccgcc
     2521 attattacga caagctatcg aattatttga tcaccaaggt gcagagccag ccttcttatt
     2581 cggccttgaa ttgatcatat gcggattaga aaaacaactt aaatgtgaaa gtgggtccgc
     2641 gtacagccgc gcgcgtacga aaaacaatta cgggtctacc atcgagggcc tgctcgatct
     2701 cccggacgac gacgcccccg aagaggcggg gctggcggct ccgcgcctgt cctttctccc
     2761 cgcgggacac acgcgcagac tgtcgacggc ccccccgacc gatgtcagcc tgggggacga
     2821 gctccactta gacggcgagg acgtggcgat ggcgcatgcc gacgcgctag acgatttcga
     2881 tctggacatg ttgggggacg gggattcccc gggtccggga tttacccccc acgactccgc
     2941 cccctacggc gctctggata tggccgactt cgagtttgag cagatgttta ccgatgccct
     3001 tggaattgac gagtacggtg ggtagtctag agggcccgtt taaacccgct gatcagcctc
     3061 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     3121 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg
     3181 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga
     3241 ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga
     3301 aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc
     3361 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     3421 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     3481 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     3541 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     3601 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     3661 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     3721 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt
     3781 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     3841 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     3901 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg
     3961 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     4021 tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt
     4081 tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct
     4141 gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt
     4201 tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc
     4261 tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag
     4321 accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct atcgtggctg
     4381 gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac
     4441 tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc
     4501 gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc
     4561 tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc
     4621 ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg
     4681 ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat
     4741 gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc
     4801 cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa
     4861 gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat
     4921 tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt
     4981 tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg
     5041 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
     5101 agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata
     5161 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     5221 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga
     5281 cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     5341 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     5401 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     5461 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     5521 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     5581 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     5641 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     5701 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     5761 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     5821 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     5881 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     5941 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     6001 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     6061 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     6121 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     6181 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     6241 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     6301 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     6361 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     6421 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     6481 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     6541 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     6601 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     6661 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     6721 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     6781 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     6841 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     6901 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     6961 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     7021 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     7081 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     7141 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     7201 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     7261 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     7321 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     7381 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     7441 ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tctcccgatc ccctatggtg
     7501 cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatctgc tccctgcttg
     7561 tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt
     7621 gaccgacaat tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt
     7681 acgggccaga tatacgcgtt
//
