LOCUS       sequence-430883-        6809 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 6809)
  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 6809)
  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: OXER1-DuET
FEATURES             Location/Qualifiers
     source          1..6809
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        86..289
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     240..260
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     334..353
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        334..352
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1871..1888)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1877..2101
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2147..2575
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2234..2253)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2444..2465
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2584..2604)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2589..2918
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2769..2904
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2831..2850
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2985..3779
                     /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(3039..3058)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3649..3668
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3953..4074
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3990..4009)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4044..4063
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4123..4139)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4123..4139)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4136..4158)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4147..4163
                     /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(4171..4177,4178..4195,4196..4201))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4216..4237
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4354..4371)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4525..5113)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4605..4624)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5284..6075,6076..6144))
                     /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     5907..5926
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6145..6249)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6324..6343)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        join(6515..6809,1..85)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta
       61 cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg
      121 atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt
      181 gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac
      241 gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctc tctggctaac
      301 tagagaaccc actgcttact ggcttatcga aattaatacg actcactata gggagaccca
      361 agctggctag cgtttaaact taagcttagc gccaccatga agacgatcat cgccctgagc
      421 tacatcttct gcctggtatt cgccgactac aaggacgatg atgacgccag catcgatatg
      481 ctttgtcatc ggggtggaca gctgatcgtt cctatcatac cactctgtcc cgagcactcc
      541 tgccgaggac ggagattgca gaacttgttg agtgggcctt ggcccaagca acctatggaa
      601 ctccacaacc tttccagccc ctctccatcc ctttccagtt ctgtgctccc gcccagtttt
      661 agtccatctc cgagtagcgc cccatccgct ttcaccacag tgggaggatc tagcggcggc
      721 ccatgccacc ccacttcatc aagcctcgtg agcgcctttc ttgcccctat ccttgccctg
      781 gagttcgtac tgggcctcgt cggtaactcc ctggccctct tcatcttctg catccatacg
      841 agaccctgga cctcaaacac agtcttcctg gttagcctgg tggcagctga ttttctgctc
      901 atttcaaacc tgcctttgcg agtggattac tatcttttgc acgagacctg gaggttcggt
      961 gcagcagcct gcaaagtcaa cctgttcatg ctgtccacca atcggacagc tagcgtggtc
     1021 tttctgaccg caattgcttt gaacagatat cttaaggtcg tacagcctca tcacgtcctt
     1081 agccgagcgt ccgttggcgc agcagcaaga gtggccggtg ggctgtgggt aggcattctt
     1141 ctccttaatg ggcacctcct tctttctacc ttcagtgggc catcctgcct gagctatcgg
     1201 gttggcacga aaccttctgc gtccctgcgc tggcaccagg ccctgtacct gctggagttt
     1261 tttctgccat tggcgctcat tttgttcgcc atcgtctcca ttgggctcac aatccgcaat
     1321 cggggtctcg ggggccaagc cgggccacag cgcgccatga gggtgctggc aatggtggtt
     1381 gccgtctaca ctatttgttt tctgcctagc atcatttttg gaatggcaag catggtggca
     1441 ttttggctgt ctgcttgtag aagtcttgat ttgtgtaccc agctgttcca tggttcattg
     1501 gctttcacct acctgaactc agttctggat cccgttctgt actgcttctc aagtcctaac
     1561 ttccttcacc agagccgcgc cctgctcggc ctgacaaggg ggcggcaggg cccagtgtcc
     1621 gatgaatcct cataccagcc cagccggcag tggaggtaca gagaggctag tcggaaggcc
     1681 gaggccatcg gcaaactgaa ggtgcagggg gaagtgagcc ttgaaaaaga aggaagtagc
     1741 caaggcatcg ataccggtgg acgcaccggt gaattcgatg aggccagcac caccgtgtct
     1801 aagaccgaga cttctcaggt ggcccctgcc taactcgagt ctagagggcc cgtttaaacc
     1861 cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
     1921 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
     1981 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac
     2041 agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg
     2101 gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc
     2161 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     2221 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     2281 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     2341 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     2401 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     2461 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     2521 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc
     2581 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     2641 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     2701 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     2761 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     2821 taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt
     2881 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat
     2941 ccattttcgg atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg
     3001 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
     3061 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
     3121 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc
     3181 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
     3241 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     3301 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     3361 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     3421 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     3481 cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg
     3541 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     3601 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     3661 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     3721 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag
     3781 cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt
     3841 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     3901 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt
     3961 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     4021 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     4081 ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt
     4141 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     4201 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     4261 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     4321 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     4381 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     4441 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     4501 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     4561 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     4621 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     4681 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     4741 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     4801 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     4861 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     4921 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacgg tatttggtat
     4981 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     5041 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     5101 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     5161 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     5221 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     5281 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     5341 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     5401 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     5461 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     5521 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     5581 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     5641 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     5701 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     5761 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     5821 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     5881 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     5941 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     6001 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     6061 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     6121 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     6181 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     6241 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatctccc
     6301 gatcccctat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat
     6361 ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg cgagcaaaat ttaagctaca
     6421 acaaggcaag gcttgaccga catttgcatg aagaatctgc ttagggttag gcgttttgcg
     6481 ctgcttcgcg atgtacgggc cagatatacg cgttgacatt gattattgac tagttattaa
     6541 tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg cgttacataa
     6601 cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata
     6661 atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag
     6721 tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc
     6781 cctattgacg tcaatgacgg taaatggcc
//
