LOCUS       sequence-430918-        6551 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 6551)
  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 6551)
  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: OXGR1-DuET
FEATURES             Location/Qualifiers
     source          1..6551
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      399..987
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     888..907
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1141..1158
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1275..1296
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1311..1316,1317..1334,1335..1341)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1349..1365
                     /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     1354..1376
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1373..1389
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1373..1389
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    1438..1559
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1449..1468)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1503..1522
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(1733..2527)
                     /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(1844..1863)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     2454..2473
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(2594..2923)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2608..2743
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2662..2681)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     2908..2928
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(2937..3365)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3047..3068)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3259..3278
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    3411..3635
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     3624..3641
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     primer_bind     complement(4901..4920)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4902..4920)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(4965..5168)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(4994..5014)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        5169..5548
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     5720..5739
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        5814..5918
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5919..5987,5988..6551,1..228)
                     /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(6137..6156)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
       61 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
      121 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
      181 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
      241 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
      301 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
      361 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
      421 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
      481 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
      541 accgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
      601 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
      661 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
      721 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
      781 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
      841 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
      901 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
      961 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     1021 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     1081 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
     1141 agcgagtcag tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tctccccgcg
     1201 cgttggccga ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt
     1261 gagcgcaacg caattaatgt gagttagctc actcattagg caccccaggc tttacacttt
     1321 atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca cacaggaaac
     1381 agctatgacc atgattacgc caagctctag ctagaggtcg acggtataca gacatgataa
     1441 gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt
     1501 gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagttg
     1561 gggtgggcga agaactccag catgagatcc ccgcgctgga ggatcatcca gccggcgtcc
     1621 cggaaaacga ttccgaagcc caacctttca tagaaggcgg cggtggaatc gaaatctcgt
     1681 gatggcaggt tgggcgtcgc ttggtcggtc atttcgaacc ccagagtccc gctcagaaga
     1741 actcgtcaag aaggcgatag aaggcgatgc gctgcgaatc gggagcggcg ataccgtaaa
     1801 gcacgaggaa gcggtcagcc cattcgccgc caagctcttc agcaatatca cgggtagcca
     1861 acgctatgtc ctgatagcgg tccgccacac ccagccggcc acagtcgatg aatccagaaa
     1921 agcggccatt ttccaccatg atattcggca agcaggcatc gccatgggtc acgacgagat
     1981 cctcgccgtc gggcatgcgc gccttgagcc tggcgaacag ttcggctggc gcgagcccct
     2041 gatgctcttc gtccagatca tcctgatcga caagaccggc ttccatccga gtacgtgctc
     2101 gctcgatgcg atgtttcgct tggtggtcga atgggcaggt agccggatca agcgtatgca
     2161 gccgccgcat tgcatcagcc atgatggata ctttctcggc aggagcaagg tgagatgaca
     2221 ggagatcctg ccccggcact tcgcccaata gcagccagtc ccttcccgct tcagtgacaa
     2281 cgtcgagcac agctgcgcaa ggaacgcccg tcgtggccag ccacgatagc cgcgctgcct
     2341 cgtcctgcag ttcattcagg gcaccggaca ggtcggtctt gacaaaaaga accgggcgcc
     2401 cctgcgctga cagccggaac acggcggcat cagagcagcc gattgtctgt tgtgcccagt
     2461 catagccgaa tagcctctcc acccaagcgg ccggagaacc tgcgtgcaat ccatcttgtt
     2521 caatcatgcg aaacgatcct catcctgtct cttgatcaga tccgaaaatg gatatacaag
     2581 ctcccgggag ctttttgcaa aagcctaggc ctccaaaaaa gcctcctcac tacttctgga
     2641 atagctcaga ggcagaggcg gcctcggcct ctgcataaat aaaaaaaatt agtcagccat
     2701 ggggcggaga atgggcggaa ctgggcggag ttaggggcgg gatgggcgga gttaggggcg
     2761 ggactatggt tgctgactaa ttgagatgca tgctttgcat acttctgcct gctggggagc
     2821 ctggggactt tccacacctg gttgctgact aattgagatg catgctttgc atacttctgc
     2881 ctgctgggga gcctggggac tttccacacc ctaactgaca cacattccac agaattaatt
     2941 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
     3001 cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa
     3061 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
     3121 cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa
     3181 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
     3241 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag
     3301 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
     3361 cgcgtgggga taccccctag agccccagct ggttctttcc gcctcagaag ccatagagcc
     3421 caccgcatcc ccagcatgcc tgctattgtc ttcccaatcc tcccccttgc tgtcctgccc
     3481 caccccaccc cccagaatag aatgacacct actcagacaa tgcgatgcaa tttcctcatt
     3541 ttattaggaa aggacagtgg gagtggcacc ttccagggtc aaggaaggca cgggggaggg
     3601 gcaaacaaca gatggctggc aactagaagg cacagtcgag gctgatcagc gggtttaaac
     3661 gggccctcta gattaggcag gggccacctg agaagtctcg gtcttagaca cggtggtgct
     3721 ggcctcatcg aattcaccgg tgcgtccacc ggtatcgatg gggttgttgc tataggaaat
     3781 cttcttcgcc tgctccagat tgccacttac cttgcaccgc acggtgctgc aaacggcctg
     3841 ctggaagtta tcagaaacca cgacatagag caacaaattg ccgaatgtgt tcagggccgc
     3901 cagaggccta gacacaatat atgcctcgtg gatctggttc tctatactac aagagattga
     3961 gagcagccga ctttcgattc gtatgactcg gagaatgtga aatggcagga agcaaacgta
     4021 gaaagcgaga agcagcagaa tggtgagtcg gcgggccttc tgcttcagac atgagtcagt
     4081 ctgcagtccg tgtgtcaggg tatggatgat agtagtataa cacaaagtga caatcaccag
     4141 cggcaggcaa aatgtagtgg cggtcagaat caggttatac cattttattg tgttcagttc
     4201 gtcggaactt gtcaggtcca agcatgcgga tctattcgtg cgattggttg acgtgatcag
     4261 aaaggtcatg gggatcaccg cgacaagtga aatgatccac accacagcac aggcgaccac
     4321 ggcacatcta gttttgtgga tgctgaaaca ggacatcggg tggatgatga cgcaatatct
     4381 gaagatgctg aagcatgtaa ggaacaaaat ggaggagtag aggttaaagt ggaaactaaa
     4441 gcgaataaat ttacacatga aatcaccgaa aatccagttt tctccgcttg cgtaataatg
     4501 aatcaggaat ggcaagctgg tcaaatacag caggtcggta caagccaggt tcaacataat
     4561 gatcgtgctg gatttccaag gtcgcatctt gaagatgtat gtggatataa ccacagcgtt
     4621 cccagggaat ccaacgagaa agatgatccc ataaatcacg ggcaggtaat gcatcttgag
     4681 gggaatattc tcatcagtgc aattcccgaa ggcagcagcg taatcaggga aatcgctggc
     4741 gttagccaga tagtccagag gctcgttcat atcgatgctg gcgtcatcat cgtccttgta
     4801 gtcggcgaat accaggcaga agatgtagct cagggcgatg atcgtcttca tggtggcgcg
     4861 gccgctaagc ttaagtttaa acgctagcca gcttgggtct ccctatagtg agtcgtatta
     4921 atttcgataa gccagtaagc agtgggttct ctagttagcc agagagctct gcttatatag
     4981 acctcccacc gtacacgcct accgcccatt tgcgtcaatg gggcggagtt gttacgacat
     5041 tttggaaagt cccgttgatt ttggtgccaa aacaaactcc cattgacgtc aatggggtgg
     5101 agacttggaa atccccgtga gtcaaaccgc tatccacgcc cattgatgta ctgccaaaac
     5161 cgcatcacca tggtaatagc gatgactaat acgtagatgt actgccaagt aggaaagtcc
     5221 cataaggtca tgtactgggc ataatgccag gcgggccatt taccgtcatt gacgtcaata
     5281 gggggcgtac ttggcatatg atacacttga tgtactgcca agtgggcagt ttaccgtaaa
     5341 tactccaccc attgacgtca atggaaagtc cctattggcg ttactatggg aacatacgtc
     5401 attattgacg tcaatgggcg ggggtcgttg ggcggtcagc caggcgggcc atttaccgta
     5461 agttatgtaa cgcggaactc catatatggg ctatgaacta atgaccccgt aattgattac
     5521 tattaataac tagtcaataa tcaatgtcaa cgcgtatatc tggcccgtac atcgcgaagc
     5581 agcgcaaaac gcctaaccct aagcagattc ttcatgcaaa tgtcggtcaa gccttgcctt
     5641 gttgtagctt aaattttgct cgcgcactac tcagcgacct ccaacacaca agcagggagc
     5701 agatactggc ttaactatgc ggcatcagag cagattgtac tgagagtgca ccatagggga
     5761 tcgggagatc tcccgatccg tcgacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     5821 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     5881 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     5941 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     6001 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     6061 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     6121 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     6181 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     6241 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     6301 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     6361 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     6421 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     6481 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     6541 ggatggaggc g
//
