LOCUS       sequence-430802-        6938 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 6938)
  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 6938)
  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: GIPR-DuET
FEATURES             Location/Qualifiers
     source          1..6938
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    18..139
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(55..74)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     109..128
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(188..204)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(188..204)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(201..223)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    212..228
                     /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(236..242,243..260,261..266))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    281..302
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(419..436)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(590..1178)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(670..689)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1349..2140,2141..2209))
                     /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     1972..1991
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2210..2314)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2389..2408)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2580..2959
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2960..3163
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3114..3134
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     3208..3227
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3208..3226
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4874..4891)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4880..5104
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      5150..5578
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5237..5256)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5447..5468
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5587..5607)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5592..5921
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5772..5907
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5834..5853
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5988..6782
                     /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(6042..6061)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6652..6671
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
ORIGIN
        1 agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata
       61 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
      121 aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt
      181 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
      241 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
      301 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
      361 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
      421 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
      481 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
      541 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
      601 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
      661 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
      721 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
      781 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
      841 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
      901 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
      961 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     1021 acactagaag aacggtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     1081 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     1141 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     1201 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     1261 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     1321 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     1381 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     1441 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     1501 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     1561 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     1621 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     1681 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     1741 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     1801 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     1861 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     1921 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     1981 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     2041 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     2101 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     2161 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     2221 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     2281 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     2341 acgtcgacgg atcgggagat ctcccgatcc cctatggtgc actctcagta caatctgctc
     2401 tgatgccgca tagttaagcc agtatctgct ccctgcttgt gtgttggagg tcgctgagta
     2461 gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg accgacattt gcatgaagaa
     2521 tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg
     2581 acattgatta ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc
     2641 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
     2701 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
     2761 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
     2821 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
     2881 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
     2941 agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg
     3001 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
     3061 gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat
     3121 gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctctggc taactagaga
     3181 acccactgct tactggctta tcgaaattaa tacgactcac tatagggaga cccaagctgg
     3241 ctagcgttta aacttaagct tagcgccacc atgaagacga tcatcgccct gagctacatc
     3301 ttctgcctgg tattcgccga ctacaaggac gatgatgacg ccagcatcga tatgaccact
     3361 tcaccgatct tgcagcttct gctgcgcctt tctctttgtg gcctgcttct ccagcgcgcc
     3421 gagacgggct ctaaaggtca gaccgctggc gagctgtatc agcggtggga gcggtaccga
     3481 cgggaatgcc aggaaactct ggccgccgcc gagcctccct cagggcttgc atgcaacggt
     3541 tcattcgata tgtacgtgtg ctgggactat gccgccccta atgcaaccgc tcgggcgtca
     3601 tgtccttggt atttgccctg gcaccatcat gtggccgcgg gttttgtgct ccgccagtgc
     3661 ggctctgatg ggcaatgggg gctgtggaga gatcacacac agtgcgaaaa tccggagaag
     3721 aacgaagctt ttctggatca gcgcctgatc ctcgaacgcc tccaggttat gtatactgtg
     3781 ggatatagct tgtcactcgc aaccttgctt ctggctctgc tgatattgtc cctctttcga
     3841 cggcttcact gcaccaggaa ctatatccac attaatttgt tcaccagctt catgctccgg
     3901 gctgccgcca ttctgtctcg ggataggctg ctgcccagac ctggaccata tctgggggat
     3961 caggcccttg cactttggaa ccaggcgttg gccgcatgtc ggacagccca aatcgtcact
     4021 cagtactgcg tgggcgcgaa ttacacctgg ctgcttgttg aaggtgtgta cctgcacagc
     4081 cttctggtcc tggtgggcgg aagtgaggaa ggtcacttca ggtactacct tctgctcggc
     4141 tggggcgctc cagctctttt cgtgatccct tgggtgatcg ttcgatatct gtatgagaac
     4201 acacagtgct gggagcggaa cgaggtcaaa gccatatggt ggataattag aacccctatc
     4261 ctgatgacca tcctgataaa ctttctgatt ttcatccgca ttctggggat cttgctgtct
     4321 aagctcagga ccaggcagat gcgctgcaga gactatcgac ttcggctcgc acgcagtaca
     4381 ctgacgttgg tgcccctcct tggcgtccat gaggtcgtgt tcgctccagt aacagaggag
     4441 caggcaagag gggcccttcg gttcgccaag ctcgggttcg aaatctttct tagtagtttc
     4501 cagggtttcc tggtttctgt gctctactgt ttcattaaca aggaagttca gtctgagatc
     4561 aggagaggct ggcaccattg cagacttaga cgctcccttg gagaggagca acgccagctg
     4621 ccagagaggg ccttccgggc actgccgtca ggctcaggcc ctggcgaggt gccaacatca
     4681 cgaggcctgt catccggcac attgcccggt cccgggaatg aggcatctcg ggaactggaa
     4741 agttactgca tcgataccgg tggacgcacc ggtgaattcg atgaggccag caccaccgtg
     4801 tctaagaccg agacttctca ggtggcccct gcctaactcg agtctagagg gcccgtttaa
     4861 acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
     4921 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
     4981 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag
     5041 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct
     5101 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt
     5161 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     5221 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     5281 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     5341 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     5401 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     5461 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     5521 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa
     5581 ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     5641 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     5701 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     5761 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     5821 gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga
     5881 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta
     5941 tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag
     6001 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
     6061 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
     6121 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag
     6181 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
     6241 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
     6301 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
     6361 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
     6421 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
     6481 tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg
     6541 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
     6601 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
     6661 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
     6721 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
     6781 gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga
     6841 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
     6901 cggctggatg atcctccagc gcggggatct catgctgg
//
