LOCUS       sequence-430907-        8102 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 8102)
  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 8102)
  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: GRM2-DuET
FEATURES             Location/Qualifiers
     source          1..8102
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(77..181)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(256..275)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        447..826
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        827..1030
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     981..1001
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1075..1094
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1075..1093
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3905..3922)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3911..4135
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4181..4609
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4268..4287)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4478..4499
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4618..4638)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4623..4952
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4803..4938
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4865..4884
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5019..5813
                     /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(5073..5092)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5683..5702
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5987..6108
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6024..6043)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6078..6097
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6157..6173)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6157..6173)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6170..6192)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6181..6197
                     /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(6205..6211,6212..6229,6230..6235))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6250..6271
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6388..6405)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6559..7147)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6639..6658)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7318..8102,1..7,8..76))
                     /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     7941..7960
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
       61 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
      121 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
      181 gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc gggagatctc ccgatcccct
      241 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt atctgctccc
      301 tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta caacaaggca
      361 aggcttgacc gacatttgca tgaagaatct gcttagggtt aggcgttttg cgctgcttcg
      421 cgatgtacgg gccagatata cgcgttgaca ttgattattg actagttatt aatagtaatc
      481 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
      541 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
      601 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
      661 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
      721 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
      781 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
      841 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
      901 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
      961 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
     1021 aagcagagct ctctggctaa ctagagaacc cactgcttac tggcttatcg aaattaatac
     1081 gactcactat agggagaccc aagctggcta gcgtttaaac ttaagcttag cggccgcgcc
     1141 accatgaaga cgatcatcgc cctgagctac atcttctgcc tggtattcgc cgactacaag
     1201 gacgatgatg acgccagcat cgatgaggga cctgccaaga aagtgctgac ccttgagggg
     1261 gacctggtcc ttggcggact gttccccgtg caccaaaaag gtggacccgc cgaagactgc
     1321 gggccggtta acgagcatcg aggcatccag cggctcgaag cgatgctctt cgctctcgat
     1381 agaattaatc gggatcccca tctgcttcca ggcgtgcggc ttggcgcaca tattctggat
     1441 tcatgctcca aagatactca tgctctcgaa caagcactcg acttcgtgcg agctagcctc
     1501 tcccgcggcg ccgatggttc aagacacatc tgccccgatg gatcctacgc cacgcacggt
     1561 gacgccccca ccgctattac gggggtgatt gggggtagct atagtgatgt tagcatacag
     1621 gtggccaatt tgctccgcct gttccagatc ccccagatta gctatgcctc aacctcagcc
     1681 aaactgtccg ataagagtag gtatgactac ttcgcccgca cagtgcctcc agatttcttt
     1741 caagccaagg ctatggcaga gatactccgc ttctttaatt ggacatacgt ttctactgta
     1801 gccagcgaag gagactacgg ggagactggc atcgaggcgt tcgaactcga agctagagcg
     1861 agaaacatat gtgtcgccac ctccgaaaag gtcgggcggg caatgtcacg cgctgccttt
     1921 gagggagtcg ttcgagccct gctgcagaag ccttccgcaa gggttgcagt cctgtttacg
     1981 cgcagtgagg atgcgcgcga attgctggcc gcttctcaac gactgaacgc atcctttaca
     2041 tgggttgcca gtgacgggtg gggcgctctg gaaagtgtgg tggccggaag tgagggagcg
     2101 gcagaggggg caattacgat cgaactggct agctacccca tcagcgattt tgcctcctac
     2161 ttccagtccc tggacccatg gaacaacagc cgcaatcctt ggttcagaga attttgggag
     2221 cagcggttta ggtgtagctt ccgacagcgg gactgtgccg ctcactcatt gagagccgtc
     2281 ccttttgagc aggagtccaa gatcatgttc gtcgtcaatg cagtttacgc catggcccac
     2341 gcgctccata atatgcaccg ggccctgtgc cctaacacca cacgcctgtg cgatgctatg
     2401 aggcctgtca acggacgaag gctgtataag gacttcgtcc tgaacgtgaa gttcgacgct
     2461 ccctttcggc ccgccgatac acataatgag gtgcgcttcg accgcttcgg ggacgggatt
     2521 ggaagataca acatcttcac gtacctgaga gccggatcag gccgctacag ataccagaaa
     2581 gtgggttact gggccgaggg cctgaccctg gacaccagtt tgatcccctg ggctagccct
     2641 tctgcagggc ccttgccggc atctcggtgc tcagagccct gtttgcagaa cgaggtaaaa
     2701 agtgttcagc ccggggaagt gtgttgttgg ctgtgcatcc cctgtcagcc atacgagtat
     2761 cgcctggatg agttcacgtg tgcagattgc ggcctcggct attggcccaa cgccagcctg
     2821 actggctgct tcgagcttcc ccaggagtat atccgatggg gcgacgcctg ggctgtaggc
     2881 cctgtgacaa tagcttgcct cggggctctg gcaaccctgt tcgtgctggg agttttcgtt
     2941 agacacaacg ccacacccgt agtcaaggcc tctggccgcg aactctgcta catcttgctg
     3001 gggggggtgt tcctttgcta ttgcatgact ttcatcttta ttgctaaacc ttcaacagcc
     3061 gtgtgcactc tcagaaggct ggggctcggc acagcgttct cagtgtgtta ctcagccttg
     3121 ttgaccaaaa ccaaccgcat tgccaggatt tttggcggtg caagagaggg cgcccagagg
     3181 ccaagattta tttcccctgc ttcacaggtg gccatatgcc tggccttgat ttccggacag
     3241 cttttgattg tcgtggcatg gctggtagtg gaggcccctg ggactggaaa ggagacagcg
     3301 cctgaacgaa gggaggtggt caccctccgc tgcaatcatc gggacgcctc tatgctcggc
     3361 agtctggctt ataatgttct tttgatcgcg ctctgcactc tttacgcgtt caagacacgg
     3421 aaatgcccag aaaacttcaa cgaagcaaaa ttcatcggct tcactatgta cacaacgtgt
     3481 attatctggt tggcattcct gcctatattt tacgtcacct ctagcgacta tcgggtacag
     3541 actaccacta tgtgtgtgag cgtgtctctg agtgggtctg ttgtgcttgg ctgtctgttt
     3601 gcaccaaagc tgcatattat cctgttccag ccccaaaaga acgtggtcag tcatcgggcc
     3661 ccgaccagca ggtttggtag tgcagccgct cgcgcttcta gctcacttgg ccaaggctct
     3721 ggcagccaat tcgtgcccac cgtgtgcaat ggccgagaag tggtagacag tacaaccagt
     3781 agtctcatcg ataccggtgg acgcaccggt gaattcgatg aggccagcac caccgtgtct
     3841 aagaccgaga cttctcaggt ggcccctgcc taatctagag ggcccgttta aacccgctga
     3901 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct
     3961 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca
     4021 tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag
     4081 ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct
     4141 gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca
     4201 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     4261 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     4321 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     4381 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     4441 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     4501 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     4561 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga
     4621 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     4681 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     4741 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     4801 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     4861 tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag
     4921 gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt
     4981 tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc
     5041 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
     5101 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
     5161 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat
     5221 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
     5281 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
     5341 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     5401 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     5461 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     5521 ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc
     5581 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     5641 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     5701 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     5761 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac
     5821 tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc
     5881 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     5941 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc
     6001 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     6061 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat
     6121 accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     6181 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     6241 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     6301 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     6361 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     6421 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     6481 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     6541 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     6601 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     6661 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     6721 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     6781 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     6841 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     6901 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     6961 gttcttgaag tggtggccta actacggcta cactagaaga acggtatttg gtatctgcgc
     7021 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     7081 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     7141 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     7201 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     7261 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     7321 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     7381 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     7441 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     7501 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     7561 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     7621 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     7681 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     7741 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     7801 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     7861 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     7921 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     7981 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     8041 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     8101 tt
//
