LOCUS       sequence-430784-        6656 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 6656)
  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 6656)
  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: CXCR5-DuET
FEATURES             Location/Qualifiers
     source          1..6656
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..48)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     83..102
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(162..178)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(162..178)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(175..197)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    186..202
                     /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(210..216,217..234,235..240))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    255..276
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(393..410)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(564..1152)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(644..663)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1323..2114,2115..2183))
                     /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     1946..1965
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2184..2288)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2363..2382)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2554..2933
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2934..3137
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3088..3108
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     3182..3201
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3182..3200
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4566..4583)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4572..4796
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4842..5270
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4929..4948)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5139..5160
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5279..5299)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5284..5613
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5464..5599
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5526..5545
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5680..6474
                     /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(5734..5753)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6344..6363
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    join(6648..6656,1..113)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
ORIGIN
        1 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
       61 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc
      121 tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg
      181 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
      241 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
      301 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
      361 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
      421 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
      481 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
      541 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
      601 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
      661 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
      721 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
      781 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
      841 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
      901 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
      961 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacggt atttggtatc
     1021 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     1081 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     1141 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     1201 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     1261 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     1321 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     1381 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     1441 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     1501 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     1561 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     1621 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     1681 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     1741 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     1801 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     1861 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     1921 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     1981 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     2041 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     2101 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     2161 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     2221 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     2281 gttccgcgca catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctcccg
     2341 atcccctatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtatc
     2401 tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa
     2461 caaggcaagg cttgaccgac atttgcatga agaatctgct tagggttagg cgttttgcgc
     2521 tgcttcgcga tgtacgggcc agatatacgc gttgacattg attattgact agttattaat
     2581 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac
     2641 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa
     2701 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
     2761 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc
     2821 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat
     2881 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc
     2941 ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc
     3001 tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa
     3061 aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg
     3121 tctatataag cagagctctc tggctaacta gagaacccac tgcttactgg cttatcgaaa
     3181 ttaatacgac tcactatagg gagacccaag ctggctagcg tttaaactta agcttagcgc
     3241 caccatgaag acgatcatcg ccctgagcta catcttctgc ctggtattcg ccgactacaa
     3301 ggacgatgat gacgccagca tcgatatgaa ctatcccctg acccttgaga tggacctgga
     3361 gaacctggaa gacttgtttt gggaattgga tcggttggac aactacaatg acacctcttt
     3421 ggtcgagaat catctgtgcc cggcgacaga gggccccttg atggcttcat tcaaggctgt
     3481 ctttgtacct gtagcctata gcctgatttt cctgcttgga gtgataggca acgtgctggt
     3541 cctggttatc ctggagcgcc accgccagac gcggtcctct acggaaacct tcctgtttca
     3601 ccttgctgtc gcagacctgc tgctcgtgtt catcctgcct ttcgccgttg cggagggtag
     3661 cgtgggttgg gttctgggga ctttcctgtg taaaacagta attgcgttgc ataaagtaaa
     3721 tttctactgc tcatctctcc tgctcgcatg tattgctgtg gatcgctacc ttgcaattgt
     3781 gcacgcggtc catgcttaca ggcaccgccg cctgctttca atccacatca catgcggaac
     3841 tatttggttg gtcgggttcc tcttggccct gcctgagatc ttgtttgcaa aggtcagcca
     3901 ggggcaccac aacaattccc tcccccgatg caccttcagt caggaaaatc aggcagagac
     3961 tcacgcctgg ttcacaagtc gctttctcta ccatgtggct ggatttttgc tgccaatgct
     4021 cgtcatgggc tggtgctatg tgggcgtcgt gcatcgcctg agacaagcac aaaggaggcc
     4081 acaacggcag aaggccgtcc gagttgccat tctggtgaca agcatctttt ttctgtgttg
     4141 gtccccttac cacattgtaa tatttttgga cacattggct cggttgaagg ccgttgacaa
     4201 cacttgtaag ctgaacggca gcctcccggt cgcgatcact atgtgcgagt ttctgggcct
     4261 ggcccattgc tgtctgaacc ccatgctgta tacatttgca ggcgtgaaat tcagaagcga
     4321 cctctctcgc ctcttgacga agctcggttg taccgggcca gctagcctgt gccagctgtt
     4381 tccaagctgg aggcggagct cattgagtga aagtgaaaat gccactagcc tgaccacctt
     4441 tatcgatacc ggtggacgca ccggtgaatt cgatgaggcc agcaccaccg tgtctaagac
     4501 cgagacttct caggtggccc ctgcctaact cgagtctaga gggcccgttt aaacccgctg
     4561 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     4621 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     4681 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     4741 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
     4801 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     4861 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     4921 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     4981 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     5041 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     5101 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     5161 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     5221 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
     5281 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     5341 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     5401 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     5461 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     5521 ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga
     5581 ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt
     5641 ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     5701 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     5761 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     5821 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta
     5881 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     5941 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     6001 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     6061 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     6121 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     6181 gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc
     6241 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     6301 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     6361 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     6421 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     6481 ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     6541 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     6601 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgttt
//
