LOCUS       sequence-430785-        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: CXCR6-DuET
FEATURES             Location/Qualifiers
     source          1..6566
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     52..71
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(282..1076)
                     /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(393..412)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1003..1022
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(1143..1472)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1157..1292
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1211..1230)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     1457..1477
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(1486..1914)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1596..1617)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1808..1827
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    1960..2184
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     2173..2190
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     primer_bind     complement(3465..3484)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3466..3484)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(3529..3732)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(3558..3578)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        3733..4112
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     4284..4303
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4378..4482
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4483..4551,4552..5343)
                     /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(4701..4720)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5514..6102
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6003..6022
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6256..6273
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6390..6411
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6426..6431,6432..6449,6450..6456)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6464..6480
                     /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     6469..6491
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6488..6504
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6488..6504
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    join(6553..6566,1..108)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(join(6564..6566,1..17))
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt
       61 gatgctattg ctttatttgt aaccattata agctgcaata aacaagttgg ggtgggcgaa
      121 gaactccagc atgagatccc cgcgctggag gatcatccag ccggcgtccc ggaaaacgat
      181 tccgaagccc aacctttcat agaaggcggc ggtggaatcg aaatctcgtg atggcaggtt
      241 gggcgtcgct tggtcggtca tttcgaaccc cagagtcccg ctcagaagaa ctcgtcaaga
      301 aggcgataga aggcgatgcg ctgcgaatcg ggagcggcga taccgtaaag cacgaggaag
      361 cggtcagccc attcgccgcc aagctcttca gcaatatcac gggtagccaa cgctatgtcc
      421 tgatagcggt ccgccacacc cagccggcca cagtcgatga atccagaaaa gcggccattt
      481 tccaccatga tattcggcaa gcaggcatcg ccatgggtca cgacgagatc ctcgccgtcg
      541 ggcatgcgcg ccttgagcct ggcgaacagt tcggctggcg cgagcccctg atgctcttcg
      601 tccagatcat cctgatcgac aagaccggct tccatccgag tacgtgctcg ctcgatgcga
      661 tgtttcgctt ggtggtcgaa tgggcaggta gccggatcaa gcgtatgcag ccgccgcatt
      721 gcatcagcca tgatggatac tttctcggca ggagcaaggt gagatgacag gagatcctgc
      781 cccggcactt cgcccaatag cagccagtcc cttcccgctt cagtgacaac gtcgagcaca
      841 gctgcgcaag gaacgcccgt cgtggccagc cacgatagcc gcgctgcctc gtcctgcagt
      901 tcattcaggg caccggacag gtcggtcttg acaaaaagaa ccgggcgccc ctgcgctgac
      961 agccggaaca cggcggcatc agagcagccg attgtctgtt gtgcccagtc atagccgaat
     1021 agcctctcca cccaagcggc cggagaacct gcgtgcaatc catcttgttc aatcatgcga
     1081 aacgatcctc atcctgtctc ttgatcagat ccgaaaatgg atatacaagc tcccgggagc
     1141 tttttgcaaa agcctaggcc tccaaaaaag cctcctcact acttctggaa tagctcagag
     1201 gcagaggcgg cctcggcctc tgcataaata aaaaaaatta gtcagccatg gggcggagaa
     1261 tgggcggaac tgggcggagt taggggcggg atgggcggag ttaggggcgg gactatggtt
     1321 gctgactaat tgagatgcat gctttgcata cttctgcctg ctggggagcc tggggacttt
     1381 ccacacctgg ttgctgacta attgagatgc atgctttgca tacttctgcc tgctggggag
     1441 cctggggact ttccacaccc taactgacac acattccaca gaattaattc gcgttaaatt
     1501 tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat
     1561 caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat
     1621 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac
     1681 tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc
     1741 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga
     1801 gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca
     1861 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtggggat
     1921 accccctaga gccccagctg gttctttccg cctcagaagc catagagccc accgcatccc
     1981 cagcatgcct gctattgtct tcccaatcct cccccttgct gtcctgcccc accccacccc
     2041 ccagaataga atgacaccta ctcagacaat gcgatgcaat ttcctcattt tattaggaaa
     2101 ggacagtggg agtggcacct tccagggtca aggaaggcac gggggagggg caaacaacag
     2161 atggctggca actagaaggc acagtcgagg ctgatcagcg ggtttaaacg ggccctctag
     2221 actcgagtta ggcaggggcc acctgagaag tctcggtctt agacacggtg gtgctggcct
     2281 catcgaattc accggtgcgt ccaccggtat cgatcagctg gaacatggag gtagcctcca
     2341 cgttatggct agcggagaac gtcttgctat tgtcttctga tgacttccat tggtgagaca
     2401 cccccaggta gggcaagcag ccgatgtctt tgaccagttt ccagaaattt ttgcgaaact
     2461 tgagagaaac aaaggcgtag agcacggggt tcaggcaggc tcggagatag gcgatcgctt
     2521 cggtaaccat gatcgtgtaa tggaaactgg tcatggcgta atactcccag tgagtagacc
     2581 tgatgaactt catcagatta aatggcatct gagtgagcag aaacactgcc atcacgagga
     2641 atataatctt cagactcctg tgtttctgaa aaccaccggc gtgcaggaga gtcttgataa
     2701 tcacggagta gcaaacaatc atggtcagca aaggaaggaa aaaaccaagg gtcatctgcg
     2761 tggccagaac cacagtggag atggcttcgt catggtatcc gcaaataagc ttatccaggt
     2821 tgaagacgtt accatagata atctgcggga ggctaaccaa aagggagatg acccatataa
     2881 ggaggcttgt cactttcccc caagtcatcc gtttggcctg ctggttatag gccttggtgg
     2941 ccttgactac cacgatgaac ctatccacag tgatacatgt caggatcaac atagaggtat
     3001 aaaagtttat cgtgtagatg ccgagcaggg acttacacat aacctgcccg aagacccact
     3061 cgtgtatccc agcgtaagcc caaaaaggga gtgtacaaac aaagaccagg tcggccaatg
     3121 ggaggtttac aagaaacacg tcggtcaagc tctgcaattt atgatagaaa atacttataa
     3181 ccagaacgag tgagttgcca acgaggccac acacaaaaac caccagatac atgcagggga
     3241 gaaagacttt ggagaactgc aggaagtctt gatgctcttc ctgagagctg tcattgaagc
     3301 ttgaaaagcc gtaatcctcg tgatagtcgt gttcggccat atcgatgctg gcgtcatcat
     3361 cgtccttgta gtcggcgaat accaggcaga agatgtagct cagggcgatg atcgtcttca
     3421 tggtggcgct aagcttaagt ttaaacgcta gccagcttgg gtctccctat agtgagtcgt
     3481 attaatttcg ataagccagt aagcagtggg ttctctagtt agccagagag ctctgcttat
     3541 atagacctcc caccgtacac gcctaccgcc catttgcgtc aatggggcgg agttgttacg
     3601 acattttgga aagtcccgtt gattttggtg ccaaaacaaa ctcccattga cgtcaatggg
     3661 gtggagactt ggaaatcccc gtgagtcaaa ccgctatcca cgcccattga tgtactgcca
     3721 aaaccgcatc accatggtaa tagcgatgac taatacgtag atgtactgcc aagtaggaaa
     3781 gtcccataag gtcatgtact gggcataatg ccaggcgggc catttaccgt cattgacgtc
     3841 aatagggggc gtacttggca tatgatacac ttgatgtact gccaagtggg cagtttaccg
     3901 taaatactcc acccattgac gtcaatggaa agtccctatt ggcgttacta tgggaacata
     3961 cgtcattatt gacgtcaatg ggcgggggtc gttgggcggt cagccaggcg ggccatttac
     4021 cgtaagttat gtaacgcgga actccatata tgggctatga actaatgacc ccgtaattga
     4081 ttactattaa taactagtca ataatcaatg tcaacgcgta tatctggccc gtacatcgcg
     4141 aagcagcgca aaacgcctaa ccctaagcag attcttcatg caaatgtcgg tcaagccttg
     4201 ccttgttgta gcttaaattt tgctcgcgca ctactcagcg acctccaaca cacaagcagg
     4261 gagcagatac tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatag
     4321 gggatcggga gatctcccga tccgtcgacg tcaggtggca cttttcgggg aaatgtgcgc
     4381 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     4441 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     4501 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     4561 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     4621 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     4681 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     4741 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     4801 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     4861 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     4921 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     4981 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     5041 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     5101 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     5161 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     5221 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     5281 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     5341 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     5401 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     5461 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     5521 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     5581 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     5641 gcgcagatac caaataccgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     5701 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     5761 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     5821 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     5881 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     5941 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     6001 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     6061 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     6121 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     6181 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     6241 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa
     6301 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
     6361 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
     6421 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
     6481 tttcacacag gaaacagcta tgaccatgat tacgccaagc tctagctaga ggtcgacggt
     6541 atacagacat gataagatac attgat
//
