LOCUS       sequence-430833-        6644 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 6644)
  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 6644)
  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: GPR62-DuET
FEATURES             Location/Qualifiers
     source          1..6644
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     68..85
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    202..223
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(238..243,244..261,262..268)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    276..292
                     /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)."
     primer_bind     281..303
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     300..316
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     300..316
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    365..486
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(376..395)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     430..449
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(660..1454)
                     /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(771..790)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1381..1400
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(1521..1850)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1535..1670
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1589..1608)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     1835..1855
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(1864..2292)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1974..1995)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2186..2205
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    2338..2562
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     2551..2568
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     primer_bind     complement(3921..3940)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3922..3940)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(3985..4188)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(4014..4034)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        4189..4568
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     4740..4759
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4834..4938
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4939..5007,5008..5799)
                     /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     complement(5157..5176)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5970..6558
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6459..6478
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac
       61 cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct
      121 ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc
      181 gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt
      241 acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac
      301 aggaaacagc tatgaccatg attacgccaa gctctagcta gaggtcgacg gtatacagac
      361 atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc
      421 tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa
      481 caagttgggg tgggcgaaga actccagcat gagatccccg cgctggagga tcatccagcc
      541 ggcgtcccgg aaaacgattc cgaagcccaa cctttcatag aaggcggcgg tggaatcgaa
      601 atctcgtgat ggcaggttgg gcgtcgcttg gtcggtcatt tcgaacccca gagtcccgct
      661 cagaagaact cgtcaagaag gcgatagaag gcgatgcgct gcgaatcggg agcggcgata
      721 ccgtaaagca cgaggaagcg gtcagcccat tcgccgccaa gctcttcagc aatatcacgg
      781 gtagccaacg ctatgtcctg atagcggtcc gccacaccca gccggccaca gtcgatgaat
      841 ccagaaaagc ggccattttc caccatgata ttcggcaagc aggcatcgcc atgggtcacg
      901 acgagatcct cgccgtcggg catgcgcgcc ttgagcctgg cgaacagttc ggctggcgcg
      961 agcccctgat gctcttcgtc cagatcatcc tgatcgacaa gaccggcttc catccgagta
     1021 cgtgctcgct cgatgcgatg tttcgcttgg tggtcgaatg ggcaggtagc cggatcaagc
     1081 gtatgcagcc gccgcattgc atcagccatg atggatactt tctcggcagg agcaaggtga
     1141 gatgacagga gatcctgccc cggcacttcg cccaatagca gccagtccct tcccgcttca
     1201 gtgacaacgt cgagcacagc tgcgcaagga acgcccgtcg tggccagcca cgatagccgc
     1261 gctgcctcgt cctgcagttc attcagggca ccggacaggt cggtcttgac aaaaagaacc
     1321 gggcgcccct gcgctgacag ccggaacacg gcggcatcag agcagccgat tgtctgttgt
     1381 gcccagtcat agccgaatag cctctccacc caagcggccg gagaacctgc gtgcaatcca
     1441 tcttgttcaa tcatgcgaaa cgatcctcat cctgtctctt gatcagatcc gaaaatggat
     1501 atacaagctc ccgggagctt tttgcaaaag cctaggcctc caaaaaagcc tcctcactac
     1561 ttctggaata gctcagaggc agaggcggcc tcggcctctg cataaataaa aaaaattagt
     1621 cagccatggg gcggagaatg ggcggaactg ggcggagtta ggggcgggat gggcggagtt
     1681 aggggcggga ctatggttgc tgactaattg agatgcatgc tttgcatact tctgcctgct
     1741 ggggagcctg gggactttcc acacctggtt gctgactaat tgagatgcat gctttgcata
     1801 cttctgcctg ctggggagcc tggggacttt ccacacccta actgacacac attccacaga
     1861 attaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg
     1921 gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt
     1981 ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct
     2041 atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt
     2101 gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa
     2161 agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc
     2221 tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc
     2281 tacagggcgc gtggggatac cccctagagc cccagctggt tctttccgcc tcagaagcca
     2341 tagagcccac cgcatcccca gcatgcctgc tattgtcttc ccaatcctcc cccttgctgt
     2401 cctgccccac cccacccccc agaatagaat gacacctact cagacaatgc gatgcaattt
     2461 cctcatttta ttaggaaagg acagtgggag tggcaccttc cagggtcaag gaaggcacgg
     2521 gggaggggca aacaacagat ggctggcaac tagaaggcac agtcgaggct gatcagcggg
     2581 tttaaacggg ccctctagac tcgagttagg caggggccac ctgagaagtc tcggtcttag
     2641 acacggtggt gctggcctca tcgaattcac cggtgcgtcc accggtatcg atgctcaggc
     2701 tgctttccgg cggcccttga taggcggggc ttcgaccgcc ggcgagctca ggggtttgct
     2761 ctggggcctc ggaagggcct actgcagggc cttctggtgg cctttgaaga cactgcaaca
     2821 aggcgcgtgg gtgccatgcc tgaggggtac atgcccggac gggcccagga agagctcgcc
     2881 ttgagagccg tcccagagcc aaccttactg gccgttgcag aagtccatac agaaacggat
     2941 gagctgcgaa agcggagtag gccacccagg tcacagctgc ttcagcttca gccgctctgg
     3001 cggcaggtgc gaggcatgca catccgtagg ggagccaaca cgctgcaaac tgacctacag
     3061 caagggcagg cgcgagggcg gctttaccgc cgggcagtcg tggccgaagg gggggcaaga
     3121 tgctcagccg agaatccaga ctatcggaat gcaaccggga gcccctagcg gggcgaggtg
     3181 gccgcagagc agccctccta gccaccacga agatccctcc ataggcaccc agaagcagca
     3241 atgctggaag tgcaaaggcc aggagtgccc acaggggtct gaagggaccg aggcctccag
     3301 caagaacgga gcatctggcg ggagccgggg ggggggctgg gggtgtgccc aacagactga
     3361 gtgcgccaag caggccggcc gctgcccaca ctgcggtcaa caccagcacg ggaggaggtc
     3421 ttgacccggg gcgcagaggg tggacgatca gcctatacct tgcaaggcca agcgcagcca
     3481 ctcccagggt acacgccggc agaagagcgg cagacaggaa gcgcgccgcc cggcaagggg
     3541 ctggacccag tcttacccga cccaggcctg gcgggggggc ggcgaggagc ccgaggggca
     3601 tgatgctcgc ggcggcgagc aggtctacga cgcacagatg ggccaggtaa agggcgtcgc
     3661 ggaggcctgg tgttctcaac accacaacca acagagcgcc gttcccgagc agggcgccga
     3721 cttccacgac agcagcgagg atgagcccaa ggcttccggc cacctcgcta gcattaaggc
     3781 cagtgctgtt ggccatatcg atgctggcgt catcatcgtc cttgtagtcg gcgaatacca
     3841 ggcagaagat gtagctcagg gcgatgatcg tcttcatggt ggcgctaagc ttaagtttaa
     3901 acgctagcca gcttgggtct ccctatagtg agtcgtatta atttcgataa gccagtaagc
     3961 agtgggttct ctagttagcc agagagctct gcttatatag acctcccacc gtacacgcct
     4021 accgcccatt tgcgtcaatg gggcggagtt gttacgacat tttggaaagt cccgttgatt
     4081 ttggtgccaa aacaaactcc cattgacgtc aatggggtgg agacttggaa atccccgtga
     4141 gtcaaaccgc tatccacgcc cattgatgta ctgccaaaac cgcatcacca tggtaatagc
     4201 gatgactaat acgtagatgt actgccaagt aggaaagtcc cataaggtca tgtactgggc
     4261 ataatgccag gcgggccatt taccgtcatt gacgtcaata gggggcgtac ttggcatatg
     4321 atacacttga tgtactgcca agtgggcagt ttaccgtaaa tactccaccc attgacgtca
     4381 atggaaagtc cctattggcg ttactatggg aacatacgtc attattgacg tcaatgggcg
     4441 ggggtcgttg ggcggtcagc caggcgggcc atttaccgta agttatgtaa cgcggaactc
     4501 catatatggg ctatgaacta atgaccccgt aattgattac tattaataac tagtcaataa
     4561 tcaatgtcaa cgcgtatatc tggcccgtac atcgcgaagc agcgcaaaac gcctaaccct
     4621 aagcagattc ttcatgcaaa tgtcggtcaa gccttgcctt gttgtagctt aaattttgct
     4681 cgcgcactac tcagcgacct ccaacacaca agcagggagc agatactggc ttaactatgc
     4741 ggcatcagag cagattgtac tgagagtgca ccatagggga tcgggagatc tcccgatccg
     4801 tcgacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     4861 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     4921 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
     4981 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
     5041 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
     5101 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
     5161 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac
     5221 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
     5281 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
     5341 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
     5401 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
     5461 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc
     5521 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
     5581 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
     5641 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc
     5701 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
     5761 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
     5821 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     5881 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     5941 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     6001 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     6061 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa taccgttctt
     6121 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     6181 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     6241 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     6301 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     6361 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     6421 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     6481 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     6541 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     6601 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attc
//
