LOCUS       sequence-430761-        6626 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 6626)
  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 6626)
  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: CCR10-DuET
FEATURES             Location/Qualifiers
     source          1..6626
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(18..809,810..878))
                     /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     641..660
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(879..983)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1058..1077)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1249..1628
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1629..1832
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1783..1803
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1877..1896
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1877..1895
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3231..3248)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3237..3461
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3507..3935
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3594..3613)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3804..3825
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3944..3964)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3949..4278
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4129..4264
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4191..4210
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4345..5139
                     /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(4399..4418)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5009..5028
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5313..5434
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5350..5369)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5404..5423
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5483..5499)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5483..5499)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5496..5518)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5507..5523
                     /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(5531..5537,5538..5555,5556..5561))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5576..5597
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5714..5731)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5885..6473)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5965..5984)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
       61 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
      121 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
      181 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
      241 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
      301 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
      361 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
      421 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
      481 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
      541 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
      601 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
      661 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
      721 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
      781 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
      841 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
      901 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
      961 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga
     1021 tcgggagatc tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat
     1081 agttaagcca gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca
     1141 aaatttaagc tacaacaagg caaggcttga ccgacatttg catgaagaat ctgcttaggg
     1201 ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgttga cattgattat
     1261 tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt
     1321 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
     1381 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
     1441 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
     1501 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc
     1561 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
     1621 ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac
     1681 ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc
     1741 aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc
     1801 gtgtacggtg ggaggtctat ataagcagag ctctctggct aactagagaa cccactgctt
     1861 actggcttat cgaaattaat acgactcact atagggagac ccaagctggc tagcgtttaa
     1921 acttaagctt agcgccacca tgaagacgat catcgccctg agctacatct tctgcctggt
     1981 attcgccgac tacaaggacg atgatgacgc cagcatcgat atgggaaccg aggcaacaga
     2041 gcaggtctct tggggtcatt actctggtga cgaggaagac gcgtacagcg ccgagcctct
     2101 ccctgagctg tgctataagg cagatgtgca ggctttttcc agagcgtttc agccgagcgt
     2161 gagcttgaca gtggcggctt tgggtctggc cggcaacgga ctggtgctcg cgacacacct
     2221 cgcagctcga agagcggcca ggtcaccaac tagtgcacac ctgttgcagc ttgccctggc
     2281 tgacctgctt ttggccctga ccctgccttt tgctgccgcc ggcgctcttc agggttggtc
     2341 cctggggtca gccacgtgcc gcaccatttc tgggctctat tccgcctctt tccacgcagg
     2401 gtttctgttc cttgcctgca ttagcgctga ccgctacgtg gccattgctc gggctttgcc
     2461 agccggacct cgaccctcca cccccggacg agcacacctg gtatccgtta tcgtctggct
     2521 gctctccctt cttctcgctc tgccagccct gttgtttagc caggatggcc agagagaggg
     2581 ccaacgacgc tgccgcctga tattcccaga gggccttacc cagaccgtca aaggcgcctc
     2641 tgccgtggcc caggtggcac tgggtttcgc tctccccctg ggtgtaatgg tggcctgcta
     2701 tgccctcctg ggaagaaccc tgctggctgc ccggggccca gaacgcagaa gagcactccg
     2761 cgttgtcgtt gcactcgtgg ccgcgtttgt ggtgctccag ctgccctatt cattggcact
     2821 tctgctggat acggccgacc tccttgccgc gcgggagcgc tcctgtcctg catccaaaag
     2881 gaaagacgta gccctgttgg tgacatccgg cctggcgctt gcccgctgcg ggttgaatcc
     2941 cgtgttgtat gcttttctgg gactgcgctt tcgccaagat ctcaggcgct tgttgagggg
     3001 tgggagttgc ccttccgggc cccaaccccg gcgagggtgc ccgcgaagac cccgactgag
     3061 ttcatgttca gctcctactg agactcatag ccttagctgg gataatatcg ataccggtgg
     3121 acgcaccggt gaattcgatg aggccagcac caccgtgtct aagaccgaga cttctcaggt
     3181 ggcccctgcc taactcgagt ctagagggcc cgtttaaacc cgctgatcag cctcgactgt
     3241 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     3301 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     3361 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     3421 agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac
     3481 cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg
     3541 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     3601 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     3661 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     3721 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     3781 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     3841 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     3901 aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta
     3961 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     4021 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     4081 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     4141 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     4201 gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     4261 ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcaa
     4321 gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg
     4381 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
     4441 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
     4501 ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg
     4561 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
     4621 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
     4681 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
     4741 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
     4801 gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc
     4861 aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
     4921 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
     4981 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt
     5041 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
     5101 cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa
     5161 tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct
     5221 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     5281 gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt
     5341 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     5401 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta
     5461 gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     5521 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     5581 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     5641 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     5701 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     5761 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     5821 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     5881 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     5941 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     6001 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     6061 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     6121 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     6181 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     6241 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     6301 ggcctaacta cggctacact agaagaacgg tatttggtat ctgcgctctg ctgaagccag
     6361 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     6421 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     6481 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     6541 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     6601 ttaaatcaat ctaaagtata tatgag
//
