LOCUS       sequence-430738-        6566 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  The DuET construct encodes the named human GPCR engineered with a 
            cleaved hemagglutinin signal peptide, a 5' FLAG and a 3' 1D4 epitope
            tag optimized for expression in mammalian cells and proteomics 
            studies..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6566)
  AUTHORS   Dahl L, Kotliar IB, Bendes A, Dodig-Crnkovic T, Fromm S, Elofsson A,
            Uhlen M, Sakmar TP, Schwenk JM
  TITLE     Multiplexed selectivity screening of anti-GPCR antibodies.
  JOURNAL   Sci Adv. 2023 May 3;9(18):eadf9297. doi: 10.1126/sciadv.adf9297. 
            Epub 2023 May 3.
  PUBMED    37134173
REFERENCE   2  (bases 1 to 6566)
  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"; journalName: "Sci Adv."; 
            date: "2023-05-3"; volume: "9(18):eadf9297. doi"; pages: " 
            10.1126/sciadv.adf9297. Epub 2023 May 3"
COMMENT     Sequence Label: TAAR8-DuET
FEATURES             Location/Qualifiers
     source          1..6566
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      241..669
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(328..347)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     538..559
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(678..698)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        683..1012
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      863..998
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     925..944
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             1079..1873
                     /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(1133..1152)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1743..1762
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    2047..2168
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2084..2103)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2138..2157
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2217..2233)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2217..2233)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2230..2252)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2241..2257
                     /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(2265..2271,2272..2289,2290..2295))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2310..2331
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2448..2465)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2619..3207)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2699..2718)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3378..4169,4170..4238))
                     /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     4001..4020
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4239..4343)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4418..4437)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        4609..4988
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4989..5192
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5143..5163
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5237..5256
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5237..5255
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6531..6548)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    join(6537..6566,1..195)
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct
       61 aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg
      121 gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg
      181 cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc
      241 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
      301 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
      361 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
      421 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
      481 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
      541 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
      601 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
      661 acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc
      721 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc
      781 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat
      841 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
      901 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga
      961 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc
     1021 gggagcttgt atatccattt tcggatctga tcaagagaca ggatgaggat cgtttcgcat
     1081 gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg
     1141 ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc
     1201 gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca
     1261 ggacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct
     1321 cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga
     1381 tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg
     1441 gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat
     1501 cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga
     1561 gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg
     1621 cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg
     1681 ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat
     1741 agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct
     1801 cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga
     1861 cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg
     1921 ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt
     1981 ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc
     2041 caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     2101 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     2161 gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca
     2221 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     2281 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     2341 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     2401 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     2461 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     2521 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     2581 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     2641 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     2701 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     2761 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     2821 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     2881 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     2941 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     3001 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     3061 acggtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     3121 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     3181 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     3241 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     3301 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     3361 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     3421 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     3481 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     3541 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     3601 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     3661 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     3721 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     3781 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     3841 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     3901 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     3961 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     4021 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     4081 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     4141 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     4201 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     4261 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     4321 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga
     4381 tcgggagatc tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat
     4441 agttaagcca gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca
     4501 aaatttaagc tacaacaagg caaggcttga ccgacatttg catgaagaat ctgcttaggg
     4561 ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgttga cattgattat
     4621 tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt
     4681 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
     4741 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
     4801 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
     4861 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc
     4921 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
     4981 ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac
     5041 ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc
     5101 aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc
     5161 gtgtacggtg ggaggtctat ataagcagag ctctctggct aactagagaa cccactgctt
     5221 actggcttat cgaaattaat acgactcact atagggagac ccaagctggc tagcgtttaa
     5281 acttaagctt agcgccacca tgaagacgat catcgccctg agctacatct tctgcctggt
     5341 attcgccgac tacaaggacg atgatgacgc cagcatcgat atgacctcta atttttccca
     5401 acccgtagtt cagctgtgtt atgaagacgt gaatggctca tgtatagaga ccccttacag
     5461 tcctgggtca cgcgtcattc tgtacactgc cttttcattt gggtctctgc tcgctgtgtt
     5521 tggcaatctg cttgtgatga ctagtgtact gcacttcaaa cagttgcaca gcccaaccaa
     5581 tttcctgatt gcatcattgg cgtgcgctga cttcctcgtg ggggtcacag tgatgttgtt
     5641 cagtatggtg cggactgttg aatcctgctg gtactttggc gccaagttct gtacccttca
     5701 cagttgttgt gacgtcgcct tctgctattc ttcagtcctg catctgtgct ttatttgtat
     5761 tgacaggtac attgtagtca cagacccgct ggtttacgct acaaaattta cagtctccgt
     5821 ctccggcatc tgcatcagtg tgtcctggat cttgcccctt acttacagcg gcgccgtgtt
     5881 ttacactggc gttaatgacg atggactgga agaattggtg agcgcactca actgtgttgg
     5941 cggctgtcaa atcatcgtga gccagggatg ggtgctgatt gactttctgc tgttcttcat
     6001 tcccactctg gtgatgatca ttctctacag caagatcttc ctcatcgcca aacagcaggc
     6061 catcaagatc gaaacaacct ccagtaaggt agagagttcc agtgagtctt ataagatccg
     6121 ggtggcgaag agagagagaa aggcagctaa gaccctgggg gtgacggtac tggcctttgt
     6181 aatttcatgg ctcccatata cagtggatat ccttatagat gcatttatgg gatttctgac
     6241 tcccgcttac atttacgaga tttgttgctg gagcgcctat tacaattctg ctatgaatcc
     6301 acttatttac gcgctcttct acccttggtt ccgaaaagca atcaaactga ttctgtcagg
     6361 cgatgtgctc aaggcaagtt ccagcacaat tagcttgttc ctggaaatcg ataccggtgg
     6421 acgcaccggt gaattcgatg aggccagcac caccgtgtct aagaccgaga cttctcaggt
     6481 ggcccctgcc taactcgagt ctagagggcc cgtttaaacc cgctgatcag cctcgactgt
     6541 gccttctagt tgccagccat ctgttg
//
