LOCUS       sequence-430861-        6977 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 6977)
  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 6977)
  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: HTR7-DuET
FEATURES             Location/Qualifiers
     source          1..6977
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    157..278
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(194..213)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     248..267
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(327..343)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(327..343)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(340..362)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    351..367
                     /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(375..381,382..399,400..405))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    420..441
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(558..575)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(729..1317)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(809..828)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1488..2279,2280..2348))
                     /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     2111..2130
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2349..2453)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2528..2547)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2719..3098
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3099..3302
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3253..3273
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     3347..3366
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3347..3365
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5052..5069)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5058..5282
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      5328..5756
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5415..5434)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5625..5646
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5765..5785)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5770..6099
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5950..6085
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6012..6031
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             6166..6960
                     /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(6220..6239)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6830..6849
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
ORIGIN
        1 gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc
       61 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
      121 cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat
      181 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
      241 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc
      301 tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
      361 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
      421 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
      481 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
      541 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
      601 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
      661 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
      721 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
      781 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
      841 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
      901 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
      961 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     1021 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     1081 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     1141 tggtggccta actacggcta cactagaaga acggtatttg gtatctgcgc tctgctgaag
     1201 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     1261 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     1321 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     1381 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     1441 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     1501 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     1561 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     1621 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     1681 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     1741 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     1801 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     1861 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     1921 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     1981 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     2041 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     2101 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     2161 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     2221 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     2281 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     2341 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     2401 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     2461 ccccgaaaag tgccacctga cgtcgacgga tcgggagatc tcccgatccc ctatggtgca
     2521 ctctcagtac aatctgctct gatgccgcat agttaagcca gtatctgctc cctgcttgtg
     2581 tgttggaggt cgctgagtag tgcgcgagca aaatttaagc tacaacaagg caaggcttga
     2641 ccgacatttg catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac
     2701 gggccagata tacgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg
     2761 ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc
     2821 cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca
     2881 tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg
     2941 cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg
     3001 acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt
     3061 ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca
     3121 tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg
     3181 tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact
     3241 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag
     3301 ctctctggct aactagagaa cccactgctt actggcttat cgaaattaat acgactcact
     3361 atagggagac ccaagctggc tagcgtttaa acttaagctt agcgccacca tgaagacgat
     3421 catcgccctg agctacatct tctgcctggt attcgccgac tacaaggacg atgatgacgc
     3481 cagcatcgat atgatggacg ttaacagctc tggaagacct gatctgtatg gccacctgcg
     3541 gagctttctc ttgcccgaag tcggcagagg cctgcctgac ctcagccctg atggaggcgc
     3601 cgaccccgtc gcaggatcct gggccccaca tctgctgtcc gaagtaacgg cgtcccctgc
     3661 tccaacctgg gatgcccccc ctgataatgc ctccgggtgc ggcgaacaga taaattatgg
     3721 acgagtggaa aaggtagtga tagggtcaat actgactctt attactcttc tgactatcgc
     3781 tggcaattgc ctcgtagtaa tctcagtttg ttttgtaaaa aaactccggc agcccagtaa
     3841 ctacctgatc gtctcactcg cactggccga tctcagtgtg gcagtcgctg ttatgccgtt
     3901 cgtctctgtc acagatctga taggtggtaa gtggatcttc ggacattttt tttgtaacgt
     3961 gtttattgcg atggacgtta tgtgttgcac cgcctcaatt atgactctct gtgtgatcag
     4021 tatcgacaga tacttgggca ttacacgccc cttgacatac ccagtgagac aaaatggaaa
     4081 gtgcatggcc aagatgattc tcagtgtatg gctgcttagc gccagcatca cccttcctcc
     4141 actctttggg tgggcacaaa atgtaaacga tgataaagtc tgtttgattt cacaagactt
     4201 tggttatacg atttattcca ccgctgtcgc cttttacatt cccatgtctg tgatgctgtt
     4261 catgtactat cagatctata aggcggctcg aaagagtgcc gccaaacata agtttccggg
     4321 gttccccaga gtggagcccg atagcgtaat cgcactgaac ggtattgtga aactgcagaa
     4381 agaagtggaa gagtgtgcga atctctctag actgcttaag catgagcgaa aaaatatctc
     4441 catcttcaag agggaacaaa aagccgccac caccctcggt atcatcgtgg gagcgttcac
     4501 ggtgtgttgg ctgcctttct tcctcctgtc cactgctaga ccattcattt gcggaacttc
     4561 ctgctcttgt attccgctgt gggttgaaag gacctttctg tggctgggtt acgctaactc
     4621 actgattaat ccattcattt acgcattttt caaccgggac ctgaggacca cctacagatc
     4681 cttgttgcag tgccagtacc ggaacataaa ccgcaaactt tcagccgccg gaatgcacga
     4741 agccctgaaa ctggccgaaa gacctgaaag gcccgaattc gtccttagag cctgtactcg
     4801 ccgcgtcttg cttaggcccg agaagaggcc tcccgtgagt gtgtgggtct tgcagagccc
     4861 agatcatcac aactggctgg ccgacaagat gcttactaca gttgagaaga aggtaatgat
     4921 ccatgacatc gataccggtg gacgcaccgg tgaattcgat gaggccagca ccaccgtgtc
     4981 taagaccgag acttctcagg tggcccctgc ctaactcgag tctagagggc ccgtttaaac
     5041 ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc
     5101 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga
     5161 aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga
     5221 cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat
     5281 ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag
     5341 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     5401 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     5461 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     5521 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     5581 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     5641 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     5701 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattaatt
     5761 ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt
     5821 atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca
     5881 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta
     5941 actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga
     6001 ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct attccagaag
     6061 tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg agcttgtata
     6121 tccattttcg gatctgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat
     6181 ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca
     6241 caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg
     6301 gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg
     6361 cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact
     6421 gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct
     6481 caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg
     6541 cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt
     6601 actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc
     6661 gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc
     6721 gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga
     6781 ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc
     6841 cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt
     6901 atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga
     6961 gcgggactct ggggttc
//
