LOCUS       sequence-430783-        6596 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 6596)
  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 6596)
  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: CXCR4-DuET
FEATURES             Location/Qualifiers
     source          1..6596
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(496..515)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(497..515)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(560..763)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(589..609)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        764..1143
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     1315..1334
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        1409..1513
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1514..1582,1583..2374)
                     /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(1732..1751)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2545..3133
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3034..3053
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3287..3304
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3421..3442
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3457..3462,3463..3480,3481..3487)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3495..3511
                     /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     3500..3522
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3519..3535
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3519..3535
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    3584..3705
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3595..3614)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3649..3668
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(3879..4673)
                     /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(3990..4009)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4600..4619
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(4740..5069)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4754..4889
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4808..4827)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     5054..5074
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(5083..5511)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5193..5214)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5405..5424
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    5557..5781
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     5770..5787
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
ORIGIN
        1 acggaactat aaagattcac agtgtagata acgtgcactg ccttacagag gaagttgccg
       61 aagtaccaat tagcgacagc gtcaactgcc caaaaaggca gggtaatcac aaacaggagg
      121 tcggccacgg acagatggag cctatacttg tcggtcattg acctcagctt cttttggtat
      181 cccatcacca ggatgaccag gccgttccct acgatgcctg tgagaaagat gatgctgtag
      241 atggtgggga gaaagatctt gttaaagttc gcgttttcct ctctaaaaca cggctctttc
      301 atagagtcgt aatcgccgga acccatctct tccgtgtaat tatctgatgt gtagattgat
      361 atcccctcca tatcgatgct ggcgtcatca tcgtccttgt agtcggcgaa taccaggcag
      421 aagatgtagc tcagggcgat gatcgtcttc atggtggcgc taagcttaag tttaaacgct
      481 agccagcttg ggtctcccta tagtgagtcg tattaatttc gataagccag taagcagtgg
      541 gttctctagt tagccagaga gctctgctta tatagacctc ccaccgtaca cgcctaccgc
      601 ccatttgcgt caatggggcg gagttgttac gacattttgg aaagtcccgt tgattttggt
      661 gccaaaacaa actcccattg acgtcaatgg ggtggagact tggaaatccc cgtgagtcaa
      721 accgctatcc acgcccattg atgtactgcc aaaaccgcat caccatggta atagcgatga
      781 ctaatacgta gatgtactgc caagtaggaa agtcccataa ggtcatgtac tgggcataat
      841 gccaggcggg ccatttaccg tcattgacgt caataggggg cgtacttggc atatgataca
      901 cttgatgtac tgccaagtgg gcagtttacc gtaaatactc cacccattga cgtcaatgga
      961 aagtccctat tggcgttact atgggaacat acgtcattat tgacgtcaat gggcgggggt
     1021 cgttgggcgg tcagccaggc gggccattta ccgtaagtta tgtaacgcgg aactccatat
     1081 atgggctatg aactaatgac cccgtaattg attactatta ataactagtc aataatcaat
     1141 gtcaacgcgt atatctggcc cgtacatcgc gaagcagcgc aaaacgccta accctaagca
     1201 gattcttcat gcaaatgtcg gtcaagcctt gccttgttgt agcttaaatt ttgctcgcgc
     1261 actactcagc gacctccaac acacaagcag ggagcagata ctggcttaac tatgcggcat
     1321 cagagcagat tgtactgaga gtgcaccata ggggatcggg agatctcccg atccgtcgac
     1381 gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     1441 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     1501 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     1561 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     1621 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     1681 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     1741 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     1801 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     1861 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     1921 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     1981 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     2041 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     2101 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     2161 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     2221 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     2281 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     2341 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
     2401 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     2461 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     2521 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     2581 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     2641 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaataccg ttcttctagt
     2701 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     2761 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     2821 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     2881 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     2941 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     3001 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     3061 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     3121 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     3181 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     3241 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     3301 cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
     3361 ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat
     3421 taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg
     3481 tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga
     3541 ttacgccaag ctctagctag aggtcgacgg tatacagaca tgataagata cattgatgag
     3601 tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat
     3661 gctattgctt tatttgtaac cattataagc tgcaataaac aagttggggt gggcgaagaa
     3721 ctccagcatg agatccccgc gctggaggat catccagccg gcgtcccgga aaacgattcc
     3781 gaagcccaac ctttcataga aggcggcggt ggaatcgaaa tctcgtgatg gcaggttggg
     3841 cgtcgcttgg tcggtcattt cgaaccccag agtcccgctc agaagaactc gtcaagaagg
     3901 cgatagaagg cgatgcgctg cgaatcggga gcggcgatac cgtaaagcac gaggaagcgg
     3961 tcagcccatt cgccgccaag ctcttcagca atatcacggg tagccaacgc tatgtcctga
     4021 tagcggtccg ccacacccag ccggccacag tcgatgaatc cagaaaagcg gccattttcc
     4081 accatgatat tcggcaagca ggcatcgcca tgggtcacga cgagatcctc gccgtcgggc
     4141 atgcgcgcct tgagcctggc gaacagttcg gctggcgcga gcccctgatg ctcttcgtcc
     4201 agatcatcct gatcgacaag accggcttcc atccgagtac gtgctcgctc gatgcgatgt
     4261 ttcgcttggt ggtcgaatgg gcaggtagcc ggatcaagcg tatgcagccg ccgcattgca
     4321 tcagccatga tggatacttt ctcggcagga gcaaggtgag atgacaggag atcctgcccc
     4381 ggcacttcgc ccaatagcag ccagtccctt cccgcttcag tgacaacgtc gagcacagct
     4441 gcgcaaggaa cgcccgtcgt ggccagccac gatagccgcg ctgcctcgtc ctgcagttca
     4501 ttcagggcac cggacaggtc ggtcttgaca aaaagaaccg ggcgcccctg cgctgacagc
     4561 cggaacacgg cggcatcaga gcagccgatt gtctgttgtg cccagtcata gccgaatagc
     4621 ctctccaccc aagcggccgg agaacctgcg tgcaatccat cttgttcaat catgcgaaac
     4681 gatcctcatc ctgtctcttg atcagatccg aaaatggata tacaagctcc cgggagcttt
     4741 ttgcaaaagc ctaggcctcc aaaaaagcct cctcactact tctggaatag ctcagaggca
     4801 gaggcggcct cggcctctgc ataaataaaa aaaattagtc agccatgggg cggagaatgg
     4861 gcggaactgg gcggagttag gggcgggatg ggcggagtta ggggcgggac tatggttgct
     4921 gactaattga gatgcatgct ttgcatactt ctgcctgctg gggagcctgg ggactttcca
     4981 cacctggttg ctgactaatt gagatgcatg ctttgcatac ttctgcctgc tggggagcct
     5041 ggggactttc cacaccctaa ctgacacaca ttccacagaa ttaattcgcg ttaaattttt
     5101 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     5161 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     5221 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     5281 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     5341 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     5401 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     5461 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tggggatacc
     5521 ccctagagcc ccagctggtt ctttccgcct cagaagccat agagcccacc gcatccccag
     5581 catgcctgct attgtcttcc caatcctccc ccttgctgtc ctgccccacc ccacccccca
     5641 gaatagaatg acacctactc agacaatgcg atgcaatttc ctcattttat taggaaagga
     5701 cagtgggagt ggcaccttcc agggtcaagg aaggcacggg ggaggggcaa acaacagatg
     5761 gctggcaact agaaggcaca gtcgaggctg atcagcgggt ttaaacgggc cctctagact
     5821 cgagttaggc aggggccacc tgagaagtct cggtcttaga cacggtggtg ctggcctcat
     5881 cgaattcacc ggtgcgtcca ccggtatcga tagagctatg aaatgagctg gattcggact
     5941 cagtgctgac actgctatgt ccgcctcgtt ttcccttgga gaggattttc agagagctcc
     6001 ccctagacac tgatgtcagt gcatgttggg cgcttgtttt aaatttggcg ccaagaaacg
     6061 cgtagagaat cggattcaaa caacagtgaa agaaggcgag ggcctcggtt atagaaatcc
     6121 acttatgaac ggtgttctcg aactcgcagc cctgcttgat aatttccagg agtataaatg
     6181 agtcaatgct gatgccgata tagtagggca gccaacatgc gaagaaggcc agtatcagga
     6241 tcaccgttgt tttcagagcc ttcctctttt ggtggccttt tgagtgtgac agcttagaaa
     6301 taatgatgca gtagcaagac agtatgacaa tgccgggcaa gatcagcccg accataatgt
     6361 gttggaactg aaataccact acccacagat cgtttgggta aaatcggtca caaatgtatc
     6421 tgtcatcggc ctcgctcaca ttagcgaaaa tgaagtctgg tattgtcaac aggagagctg
     6481 gaatccacac accaacgtaa accaccttct cagccaggag ttttctaggt cgttgtgagt
     6541 tggtagcgtg cacaatcgcc aaatacctgt cgagactgat aaaggccaag atcaag
//
