LOCUS       sequence-430859-        6914 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 6914)
  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 6914)
  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: HTR2C-DuET
FEATURES             Location/Qualifiers
     source          1..6914
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(548..1339,1340..1408))
                     /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     1171..1190
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1409..1513)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1588..1607)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1779..2158
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2159..2362
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2313..2333
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2407..2426
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2407..2425
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4049..4066)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4055..4279
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4325..4753
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4412..4431)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4622..4643
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4762..4782)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4767..5096
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4947..5082
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5009..5028
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5163..5957
                     /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(5217..5236)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5827..5846
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6131..6252
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6168..6187)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6222..6241
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6301..6317)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6301..6317)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6314..6336)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6325..6341
                     /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(6349..6355,6356..6373,6374..6379))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6394..6415
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6532..6549)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(6703..6914,1..377))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6783..6802)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
       61 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
      121 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
      181 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acggtatttg
      241 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
      301 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
      361 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
      421 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
      481 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
      541 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
      601 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
      661 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
      721 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
      781 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
      841 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
      901 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
      961 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     1021 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     1081 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     1141 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     1201 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     1261 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     1321 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     1381 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     1441 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     1501 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc
     1561 tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     1621 gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc
     1681 tacaacaagg caaggcttga ccgacatttg catgaagaat ctgcttaggg ttaggcgttt
     1741 tgcgctgctt cgcgatgtac gggccagata tacgcgttga cattgattat tgactagtta
     1801 ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac
     1861 ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc
     1921 aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt
     1981 ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac
     2041 gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac
     2101 cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt
     2161 gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac ggggatttcc
     2221 aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt
     2281 tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg
     2341 ggaggtctat ataagcagag ctctctggct aactagagaa cccactgctt actggcttat
     2401 cgaaattaat acgactcact atagggagac ccaagctggc tagcgtttaa acttaagctt
     2461 agcgccacca tgaagacgat catcgccctg agctacatct tctgcctggt attcgccgac
     2521 tacaaggacg atgatgacgc cagcatcgat atggttaatc ttcgaaacgc ggtccattca
     2581 ttcctcgtcc acctgattgg tctcctggtg tggcagtgtg atatcagtgt ttcccccgtg
     2641 gctgctatcg ttaccgatat ctttaacact tctgacggtg gaagattcaa attccctgac
     2701 ggagtgcaga actggcccgc tcttagtatt gtcatcatta tcattatgac gatcggaggt
     2761 aacatcctgg tcattatggc agtgtcaatg gaaaaaaaac ttcacaatgc gacaaactat
     2821 ttcctgatgt ccctcgccat cgccgacatg ctcgtgggac tgctggtcat gccgctgtcc
     2881 ctcctggcaa tcctgtacga ttacgtgtgg ccgttgcccc gctatctgtg cccggtctgg
     2941 atatccctgg acgtgttgtt ttctaccgca agtatcatgc acctgtgtgc aattagcctg
     3001 gatagatacg tggcaattag aaaccctatt gaacacagca ggttcaacag cagaaccaaa
     3061 gccatcatga agatcgcgat tgtgtgggcc atatctatcg gtgtctcagt gcccatcccc
     3121 gtaatcggcc tgagggacga agagaaagtc tttgttaaca acacaacctg cgtgttgaac
     3181 gaccccaact tcgtgctgat tggatctttc gtggcctttt ttattccgct cacaattatg
     3241 gtgattacct actgcctcac aatatacgtt ttgaggcgac aggccctcat gcttctccat
     3301 gggcacaccg aggagccccc cgggctctct ctggatttcc ttaagtgttg caagcggaac
     3361 acagcagaag aggagaacag cgccaaccct aaccaggacc aaaacgcccg ccgacggaaa
     3421 aagaaggaga gacgccccag gggcactatg caggctatca ataacgagcg aaaagcatct
     3481 aaagttttgg gaattgtgtt ttttgtgttt ctgattatgt ggtgcccgtt cttcatcacc
     3541 aacatcctta gcgttctctg tgagaagagc tgtaatcaga agctcatgga gaagctcctc
     3601 aacgtgtttg tctggattgg ttacgtttgc agcgggatca accctctggt gtacaccctc
     3661 ttcaacaaaa tttacagacg ggcatttagc aattatttga ggtgcaacta caaagtggaa
     3721 aaaaaacccc ctgtgcgcca gatcccaaga gtcgccgcga ccgctttgtc tggcagagag
     3781 ctcaatgtga acatctaccg gcacacaaac gaaccagtca ttgaaaaagc ctcagataat
     3841 gaaccaggga tcgaaatgca ggtggagaat ttggaactgc cagtgaaccc ctctagcgtg
     3901 gttagtgagc ggatatcatc agtgatcgat accggtggac gcaccggtga attcgatgag
     3961 gccagcacca ccgtgtctaa gaccgagact tctcaggtgg cccctgccta actcgagtct
     4021 agagggcccg tttaaacccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct
     4081 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt
     4141 tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg
     4201 ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg
     4261 gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat
     4321 ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     4381 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
     4441 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     4501 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
     4561 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     4621 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     4681 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     4741 tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     4801 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
     4861 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     4921 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     4981 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct
     5041 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
     5101 tcccgggagc ttgtatatcc attttcggat ctgatcaaga gacaggatga ggatcgtttc
     5161 gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat
     5221 tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt
     5281 cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac
     5341 tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg
     5401 tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc
     5461 aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa
     5521 tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc
     5581 gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg
     5641 aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg
     5701 acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa
     5761 atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg
     5821 acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct
     5881 tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc
     5941 ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa
     6001 cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat
     6061 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt
     6121 cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     6181 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     6241 caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg
     6301 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     6361 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     6421 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     6481 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     6541 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     6601 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     6661 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     6721 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     6781 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     6841 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     6901 ctcacgctgt aggt
//
