LOCUS       sequence-430917-        6794 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 6794)
  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 6794)
  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: NTSR1-DuET
FEATURES             Location/Qualifiers
     source          1..6794
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     276..295
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(514..618)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(693..712)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        884..1263
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1264..1467
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1418..1438
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1512..1531
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1512..1530
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3034..3051)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3040..3264
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3310..3738
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3397..3416)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3607..3628
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3747..3767)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3752..4081
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3932..4067
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3994..4013
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4148..4942
                     /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(4202..4221)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4812..4831
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5116..5237
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5153..5172)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5207..5226
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5286..5302)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5286..5302)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5299..5321)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5310..5326
                     /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(5334..5340,5341..5358,5359..5364))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5379..5400
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5517..5534)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5688..6276)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5768..5787)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6447..6794,1..444,445..513))
                     /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"
ORIGIN
        1 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
       61 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
      121 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
      181 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
      241 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
      301 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
      361 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
      421 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
      481 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
      541 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
      601 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg acggatcggg
      661 agatctcccg atcccctatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
      721 agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt
      781 taagctacaa caaggcaagg cttgaccgac atttgcatga agaatctgct tagggttagg
      841 cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc gttgacattg attattgact
      901 agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc
      961 gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg
     1021 acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa
     1081 tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca
     1141 agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac
     1201 atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc
     1261 atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga
     1321 tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg
     1381 gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta
     1441 cggtgggagg tctatataag cagagctctc tggctaacta gagaacccac tgcttactgg
     1501 cttatcgaaa ttaatacgac tcactatagg gagacccaag ctggctagcg tttaaactta
     1561 agcttagcgg ccgcgccacc atgaagacga tcatcgccct gagctacatc ttctgcctgg
     1621 tattcgccga ctacaaggac gatgatgacg ccagcatcga tatgagactg aactcttccg
     1681 ctccaggaac gccgggcaca ccggctgcag atcccttcca aagggcccag gccggcttgg
     1741 aagaggccct gctcgctccg ggcttcggaa acgcctctgg aaatgcctct gaacgcgtgc
     1801 ttgccgcacc ctccagtgag ctcgatgtta acaccgacat ctacagcaaa gtcctggtga
     1861 ccgcggtata tctggcactc ttcgtcgttg gcactgtagg taacacggtg actgctttta
     1921 cccttgcgcg gaagaagagc cttcagagcc tgcaaagcac agtacattac catctgggtt
     1981 ccctggccct ttctgacctg ctcacactcc tgctggctat gccagtggaa ctgtacaact
     2041 tcatctgggt gcaccatcct tgggcatttg gggacgctgg ctgtcgcggg tattattttc
     2101 ttagagacgc ctgcacttat gccaccgcct tgaatgtagc ctctctcagc gtagagagat
     2161 acctcgctat ctgccatccc ttcaaggcta agaccttgat gagccgaagc cgaactaaaa
     2221 agttcatctc agcaatttgg cttgcgagcg cactgctggc cgttcctatg ctgtttacta
     2281 tgggtgagca gaatagaagc gcagatggac aacacgccgg aggcctggtg tgcacaccca
     2341 ccattcacac tgccacggtg aaagtggtca ttcaggtgaa cacattcatg agctttatct
     2401 ttcccatggt cgtaatcagc gttctcaaca ccataattgc aaataagctg acagtgatgg
     2461 tgcggcaggc cgctgagcaa ggacaggtat gtaccgttgg cggcgagcac tcaacattca
     2521 gcatggcaat tgaacccggg agagtccagg cactgagaca tggagtgcgg gttctgcggg
     2581 cggtggtgat cgcgttcgta gtttgttggc tcccgtacca cgtccggcgc ctcatgttct
     2641 gctatatttc cgacgaacag tggacacctt tcctttacga cttttaccat tacttctata
     2701 tggtgacgaa cgctcttttc tatgtttcca gtacaatcaa tcctatcctt tacaacctgg
     2761 tcagtgcaaa cttccgacat atcttcctgg caacactcgc ctgtctgtgc cccgtttggc
     2821 gccgccggag gaagcgcccc gccttctcac ggaaggccga ctcagtatcc tctaaccata
     2881 ccctgagttc aaacgccact cgggaaacgc tgtacatcga taccggtgga cgcaccggtg
     2941 aattcgatga ggccagcacc accgtgtcta agaccgagac ttctcaggtg gcccctgcct
     3001 aatctagagg gcccgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc
     3061 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg
     3121 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc
     3181 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg
     3241 ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg
     3301 ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     3361 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     3421 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     3481 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     3541 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     3601 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt
     3661 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     3721 aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     3781 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     3841 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     3901 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     3961 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     4021 tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     4081 aaagctcccg ggagcttgta tatccatttt cggatctgat caagagacag gatgaggatc
     4141 gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag
     4201 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg
     4261 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa
     4321 tgaactgcag gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc
     4381 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc
     4441 ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga
     4501 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa
     4561 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct
     4621 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat
     4681 gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt
     4741 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta
     4801 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga
     4861 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg
     4921 ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg
     4981 cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     5041 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     5101 ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     5161 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     5221 ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa
     5281 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     5341 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
     5401 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     5461 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     5521 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     5581 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     5641 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     5701 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     5761 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     5821 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     5881 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     5941 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     6001 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     6061 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     6121 actagaagaa cggtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     6181 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     6241 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     6301 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     6361 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     6421 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     6481 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     6541 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     6601 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     6661 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     6721 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     6781 cgctcgtcgt ttgg
//
