LOCUS       sequence-430790-        6797 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 6797)
  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 6797)
  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: DRD4-DuET
FEATURES             Location/Qualifiers
     source          1..6797
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(378..1169,1170..1238))
                     /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     1001..1020
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1239..1343)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1418..1437)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1609..1988
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1989..2192
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2143..2163
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2237..2256
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2237..2255
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3762..3779)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3768..3992
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4038..4466
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4125..4144)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4335..4356
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4475..4495)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4480..4809
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4660..4795
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4722..4741
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4876..5670
                     /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(4930..4949)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5540..5559
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5844..5965
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5881..5900)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5935..5954
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6014..6030)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6014..6030)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6027..6049)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6038..6054
                     /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(6062..6068,6069..6086,6087..6092))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6107..6128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6245..6262)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(6416..6797,1..207))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6496..6515)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
       61 acggtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
      121 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
      181 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
      241 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
      301 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
      361 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
      421 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
      481 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
      541 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
      601 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
      661 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
      721 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
      781 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
      841 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
      901 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
      961 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     1021 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     1081 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     1141 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     1201 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     1261 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     1321 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga
     1381 tcgggagatc tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat
     1441 agttaagcca gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca
     1501 aaatttaagc tacaacaagg caaggcttga ccgacatttg catgaagaat ctgcttaggg
     1561 ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgttga cattgattat
     1621 tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt
     1681 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
     1741 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
     1801 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
     1861 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc
     1921 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
     1981 ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac
     2041 ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc
     2101 aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc
     2161 gtgtacggtg ggaggtctat ataagcagag ctctctggct aactagagaa cccactgctt
     2221 actggcttat cgaaattaat acgactcact atagggagac ccaagctggc tagcgtttaa
     2281 acttaagctt agcgccacca tgaagacgat catcgccctg agctacatct tctgcctggt
     2341 attcgccgac tacaaggacg atgatgacgc cagcatcgat atgggtaatc gaagcactgc
     2401 tgacgcagac ggcctgcttg ccgggagagg tccggccgcc ggtgccagcg ctggtgcctc
     2461 cgccggcctc gctgggcaag gggccgccgc attggtaggt ggcgtgctgc tcattggagc
     2521 tgttctggca ggaaatagtc tggtgtgtgt ctccgttgcc acagaaaggg cactccagac
     2581 tcctaccaat tcctttatcg tttctctggc agccgccgac cttcttctgg ctctccttgt
     2641 gctcccactt tttgtgtatt ctgaagtgca gggcggcgcc tggctgctct cacccagatt
     2701 gtgtgatgcc ctcatggcaa tggacgttat gctctgcacc gcttccatct ttaatctgtg
     2761 tgccatatct gtggaccgct ttgtagcagt tgccgtcccc ctgcggtaca atcgccaggg
     2821 cgggtctcgg agacagttgc tgctgatcgg cgctacttgg cttctgtccg ctgccgtggc
     2881 ggcacccgtg ctttgtggcc tgaacgacgt gcggggtcgc gatcctgccg tgtgcaggct
     2941 ggaggatagg gattatgtgg tttacagttc cgtgtgttcc ttctttctgc catgtccgct
     3001 gatgttgctg ttgtactggg ccactttccg ggggctccag cgctgggagg tcgcgagaag
     3061 ggccaagttg catggccggg ctccacgccg cccatctggt ccagggcctc cttcacccac
     3121 cccacccgcg ccgagattgc ctcaagaccc atgtggaccg gactgtgcgc cacctgcgcc
     3181 cggtctgcct cggggcccat gtggaccaga ttgtgctccg gccgccccct ctcttcccca
     3241 ggacccatgc ggccccgact gcgcccctcc cgcccccggc ctgcctcccg acccatgtgg
     3301 gtctaactgt gccccacctg atgccgtcag ggcagcagca ctgccccccc agacacctcc
     3361 ccagacacga aggagaagac gcgctaagat cactggacgc gagcgcaagg ctatgcgcgt
     3421 cttgcctgtc gttgtgggcg ccttccttct ctgctggaca ccattcttcg ttgtgcacat
     3481 cacacaggca ctttgcccag cctgctctgt ccccccgaga ctggtgtccg ctgttacgtg
     3541 gctgggatac gttaatagcg cgctgaatcc agtgatttac actgtgttta atgcagaatt
     3601 tcgcaatgtg tttaggaagg ctctccgggc ttgctgtatc gataccggtg gacgcaccgg
     3661 tgaattcgat gaggccagca ccaccgtgtc taagaccgag acttctcagg tggcccctgc
     3721 ctaactcgag tctagagggc ccgtttaaac ccgctgatca gcctcgactg tgccttctag
     3781 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     3841 tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca
     3901 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag
     3961 caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg
     4021 ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     4081 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
     4141 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     4201 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
     4261 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     4321 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
     4381 ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct
     4441 gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga
     4501 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     4561 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     4621 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     4681 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     4741 gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     4801 ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca agagacagga
     4861 tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg
     4921 gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc
     4981 gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt
     5041 gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt
     5101 ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc
     5161 gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc
     5221 atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac
     5281 caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag
     5341 gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag
     5401 gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat
     5461 atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg
     5521 gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa
     5581 tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc
     5641 ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc
     5701 aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt
     5761 tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca
     5821 tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa
     5881 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     5941 tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct
     6001 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     6061 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     6121 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     6181 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     6241 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     6301 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     6361 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     6421 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     6481 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     6541 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     6601 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     6661 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     6721 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     6781 ggattagcag agcgagg
//
