LOCUS       sequence-430905-        7127 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 7127)
  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 7127)
  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: GPR97-DuET
FEATURES             Location/Qualifiers
     source          1..7127
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     187..206
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(425..529)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(604..623)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        795..1174
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1175..1378
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1329..1349
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1423..1442
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1423..1441
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3278..3295)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3284..3508
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3554..3982
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3641..3660)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3851..3872
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3991..4011)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3996..4325
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4176..4311
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4238..4257
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4392..5186
                     /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(4446..4465)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5056..5075
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5360..5481
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5397..5416)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5451..5470
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5530..5546)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5530..5546)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5543..5565)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5554..5570
                     /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(5578..5584,5585..5602,5603..5608))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5623..5644
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5761..5778)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5932..6520)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6012..6031)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6691..7127,1..355,356..424))
                     /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"
ORIGIN
        1 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
       61 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
      121 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
      181 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
      241 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
      301 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
      361 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
      421 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
      481 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
      541 gaaaagtgcc acctgacgtc gacggatcgg gagatctccc gatcccctat ggtgcactct
      601 cagtacaatc tgctctgatg ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt
      661 ggaggtcgct gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag gcttgaccga
      721 catttgcatg aagaatctgc ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc
      781 cagatatacg cgttgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc
      841 attagttcat agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc
      901 tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt
      961 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca
     1021 cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg
     1081 taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca
     1141 gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa
     1201 tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa
     1261 tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc
     1321 cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctct
     1381 ctggctaact agagaaccca ctgcttactg gcttatcgaa attaatacga ctcactatag
     1441 ggagacccaa gctggctagc gtttaaactt aagcttagcg gccgcgccac catgaagacg
     1501 atcatcgccc tgagctacat cttctgcctg gtattcgccg actacaagga cgatgatgac
     1561 gccagcatcg atcaggagaa gccaaccgag ggacccagga acacctgtct cgggtcaaat
     1621 aacatgtacg atatctttaa tctgaatgac aaggcccttt gtttcaccaa gtgtagacaa
     1681 tctgggtccg actcctgtaa cgtggagaac ttgcagagat actggcttaa ctatgaggct
     1741 catctcatga aagagggtct cacccagaag gtgaatacac cttttctgaa agccctggtg
     1801 cagaacctgt ctacaaacac cgctgaggac ttctatttta gtctggagcc cagtcaggtc
     1861 ccacgccagg tgatgaagga tgaggacaaa ccaccagacc gcgtcagact gcccaagtca
     1921 ctcttccgat ccctcccagg gaaccgcagc gttgtgcgat tggcggttac aattctcgat
     1981 atcggcccgg ggactctgtt taaagggcct cgcctgggac tgggcgacgg aagtggtgtg
     2041 cttaataatc gcctggtggg actttccgtc gggcagatgc acgtaactaa gcttgcagaa
     2101 ccactggaaa tcgtattctc acatcagcgg cctcctccta atatgacact gacatgtgtg
     2161 ttctgggacg tcaccaaggg aacaaccgga gactggagta gtgagggctg cagcactgaa
     2221 gtacgacctg aggggacggt ttgttgttgc gaccacttga ccttcttcgc ccttctgttg
     2281 cgacctacct tggaccagag cacagtccac atcctgaccc gaatcagtca ggccggctgt
     2341 ggtgtgagca tgatttttct tgcctttaca ataatcctgt atgcctttct caggctgagc
     2401 agggaacgat ttaaatcaga agacgcccca aagatccatg ttgcactcgg aggcagcctg
     2461 ttcctcctca acttggcctt cttggtcaac gtgggctctg gcagcaaggg ttcagatgca
     2521 gcatgctggg cccgaggtgc agtgttccac tactttctgc tttgtgcctt cacctggatg
     2581 ggccttgaag cgtttcacct ctacttgttg gcagtgagag tttttaatac gtacttcgga
     2641 cactattttc tcaagctgtc cctggtgggg tggggactcc cagccttgat ggtcattggc
     2701 accggatctg ccaactctta cggactctat accatcaggg acagagaaaa ccgcaccagt
     2761 ctggagctgt gctggtttag agaggggact actatgtacg ccctttacat caccgttcac
     2821 ggttactttc tgatcacctt tctcttcgga atggtcgtac tggccctggt agtctggaag
     2881 atttttactc tgagtcgagc aactgctgtc aaggagcggg ggaagaacag gaaaaaagtg
     2941 ctcactctgc tgggactgag tagcttggtt ggggtgacat ggggccttgc tatctttaca
     3001 cctctggggc ttagtactgt gtacatcttc gctctcttca actccctgca gggcgtgttt
     3061 atttgttgtt ggtttactat tttgtacctt ccgagtcagt caactacagt gagttcctcc
     3121 acagcgaggt tggaccaagc gcatagcgca agccaagaga tcgataccgg tggacgcacc
     3181 ggtgaattcg atgaggccag caccaccgtg tctaagaccg agacttctca ggtggcccct
     3241 gcctaatcta gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc
     3301 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     3361 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     3421 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     3481 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct
     3541 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     3601 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     3661 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     3721 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     3781 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     3841 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
     3901 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     3961 taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt
     4021 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     4081 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     4141 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     4201 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     4261 cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     4321 gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag acaggatgag
     4381 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     4441 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     4501 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     4561 tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
     4621 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     4681 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     4741 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     4801 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     4861 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc
     4921 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     4981 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     5041 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
     5101 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     5161 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc
     5221 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     5281 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     5341 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa
     5401 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     5461 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc
     5521 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     5581 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     5641 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     5701 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     5761 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     5821 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     5881 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     5941 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     6001 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     6061 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     6121 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     6181 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     6241 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     6301 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     6361 ctacactaga agaacggtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     6421 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     6481 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     6541 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     6601 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     6661 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     6721 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     6781 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     6841 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     6901 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     6961 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     7021 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     7081 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtc
//
