LOCUS       sequence-430781-        6620 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 6620)
  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 6620)
  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: CXCR2-DuET
FEATURES             Location/Qualifiers
     source          1..6620
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     555..574
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(793..897)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(972..991)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1163..1542
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1543..1746
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1697..1717
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1791..1810
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1791..1809
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3139..3156)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3145..3369
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3415..3843
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3502..3521)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3712..3733
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3852..3872)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3857..4186
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4037..4172
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4099..4118
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4253..5047
                     /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(4307..4326)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4917..4936
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5221..5342
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5258..5277)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5312..5331
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5391..5407)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5391..5407)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5404..5426)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5415..5431
                     /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(5439..5445,5446..5463,5464..5469))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5484..5505
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5622..5639)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5793..6381)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5873..5892)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6552..6620,1..723,724..792))
                     /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"
ORIGIN
        1 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
       61 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
      121 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
      181 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
      241 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
      301 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
      361 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
      421 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
      481 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
      541 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
      601 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
      661 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
      721 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
      781 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
      841 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
      901 atttccccga aaagtgccac ctgacgtcga cggatcggga gatctcccga tcccctatgg
      961 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct
     1021 tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc
     1081 ttgaccgaca tttgcatgaa gaatctgctt agggttaggc gttttgcgct gcttcgcgat
     1141 gtacgggcca gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt
     1201 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
     1261 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     1321 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     1381 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
     1441 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
     1501 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
     1561 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
     1621 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac
     1681 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
     1741 agagctctct ggctaactag agaacccact gcttactggc ttatcgaaat taatacgact
     1801 cactataggg agacccaagc tggctagcgt ttaaacttaa gcttagcgcc accatgaaga
     1861 cgatcatcgc cctgagctac atcttctgcc tggtattcgc cgactacaag gacgatgatg
     1921 acgccagcat cgatatggag gacttcaaca tggagtcaga ctccttcgaa gacttctgga
     1981 aaggtgagga cctgtcaaac tattcttata gtagcacact ccctccattt cttttggacg
     2041 ctgcaccctg cgagccagaa tccctggaga ttaataagta ctttgtggtt atcatatacg
     2101 cactcgtgtt tttgctgtca ctgctcggga acagccttgt catgcttgtg attctctact
     2161 ctcgggtggg ccggtcagtt accgatgtgt atcttctgaa cctggcactt gcggatttgc
     2221 tcttcgctct cactctcccc atctgggcag cctccaaggt taacggctgg atctttggca
     2281 ccttcctttg caaggtcgtc tcactgctga aagaagtgaa tttctactca ggcattctgc
     2341 tgcttgcctg catcagcgtg gacagatatc tggcgatcgt gcacgccaca cggacactga
     2401 ctcaaaagag gtatctcgtg aaattcatat gcctttccat ttgggggctg tcactgctgc
     2461 tcgccttgcc tgtcttgttg tttagacgga ccgtatatag ctccaatgtg agccccgcat
     2521 gctatgagga tatgggcaat aatacagcca actggcgaat gctgctcaga atcctgcccc
     2581 agtcattcgg atttatcgtt cccctgctca ttatgctttt ctgttacggg tttactctgc
     2641 ggaccctttt caaagctcat atgggccaga agcacagagc aatgcgggtt atcttcgctg
     2701 tggtcctgat atttctgctg tgttggctgc cctacaacct cgtcctgctc gccgacactc
     2761 tgatgcggac tcaggtgatt caggagacat gcgagcggcg caaccacatc gacagagccc
     2821 tggacgctac agaaatactt ggcattctcc attcttgttt gaacccactg atatacgcct
     2881 ttataggaca aaagtttagg cacgggctgc tgaaaatcct cgccatccat ggccttatct
     2941 ccaaggattc cctccccaag gactcccgcc caagcttcgt gggttcatcc tccggccaca
     3001 cctccacgac cctgatcgat accggtggac gcaccggtga attcgatgag gccagcacca
     3061 ccgtgtctaa gaccgagact tctcaggtgg cccctgccta actcgagtct agagggcccg
     3121 tttaaacccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc
     3181 cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa
     3241 atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg
     3301 ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg
     3361 gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat ccccacgcgc
     3421 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     3481 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     3541 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     3601 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc
     3661 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     3721 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     3781 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     3841 attaattctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg
     3901 cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg
     3961 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc
     4021 gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca
     4081 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct ctgagctatt
     4141 ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tcccgggagc
     4201 ttgtatatcc attttcggat ctgatcaaga gacaggatga ggatcgtttc gcatgattga
     4261 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga
     4321 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg
     4381 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaggacga
     4441 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt
     4501 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct
     4561 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct
     4621 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg
     4681 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca
     4741 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg acggcgagga
     4801 tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt
     4861 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt
     4921 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct
     4981 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt
     5041 cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca
     5101 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
     5161 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc
     5221 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     5281 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     5341 tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg
     5401 tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata
     5461 aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca
     5521 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     5581 gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg
     5641 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     5701 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     5761 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     5821 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     5881 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     5941 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     6001 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     6061 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     6121 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     6181 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacggta
     6241 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     6301 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     6361 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     6421 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     6481 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     6541 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     6601 cgttcatcca tagttgcctg
//
