LOCUS       sequence-430837-        6728 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 6728)
  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 6728)
  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: GPR84-DuET
FEATURES             Location/Qualifiers
     source          1..6728
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     501..520
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(739..843)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(918..937)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1109..1488
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1489..1692
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1643..1663
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1737..1756
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1737..1755
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3193..3210)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3199..3423
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3469..3897
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3556..3575)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3766..3787
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3906..3926)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3911..4240
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4091..4226
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4153..4172
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4307..5101
                     /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(4361..4380)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4971..4990
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5275..5396
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5312..5331)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5366..5385
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5445..5461)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5445..5461)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5458..5480)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5469..5485
                     /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(5493..5499,5500..5517,5518..5523))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5538..5559
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5676..5693)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5847..6435)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5927..5946)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6606..6728,1..669,670..738))
                     /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 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
       61 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
      121 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
      181 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
      241 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
      301 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
      361 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
      421 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
      481 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
      541 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
      601 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
      661 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
      721 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
      781 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
      841 gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc tcccgatccc
      901 ctatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gtatctgctc
      961 cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc tacaacaagg
     1021 caaggcttga ccgacatttg catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt
     1081 cgcgatgtac gggccagata tacgcgttga cattgattat tgactagtta ttaatagtaa
     1141 tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg
     1201 gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg
     1261 tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta
     1321 cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt
     1381 gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac
     1441 tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt
     1501 tggcagtaca tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac
     1561 cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt
     1621 cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat
     1681 ataagcagag ctctctggct aactagagaa cccactgctt actggcttat cgaaattaat
     1741 acgactcact atagggagac ccaagctggc tagcgtttaa acttaagctt agcgccacca
     1801 tgaagacgat catcgccctg agctacatct tctgcctggt attcgccgac tacaaggacg
     1861 atgatgacgc cagcatcgat atgtggaatt cttcagacgc caattttagc tgctaccacg
     1921 aaagcgtgct ggggtaccga tatgtcgccg tttcttgggg cgtggtggtc gccgtgacag
     1981 gtaccgttgg gaatgttctc accttgttgg cactggcaat ccagccgaag ctgagaacaa
     2041 ggttcaacct gcttatagcc aacctcactc tggctgatct gctgtactgc actctgctgc
     2101 agcctttttc tgtggacaca tacctgcatt tgcactggcg gacaggagcc acattttgcc
     2161 gggtgttcgg cctcttgctc ttcgcctcca actccgtgtc catcttgaca ctttgtctta
     2221 tcgcattggg gcgctacctg cttattgccc accccaagct gttcccccaa gtcttctctg
     2281 ccaaaggaat cgtcctggca cttgtgtcaa cttgggtcgt gggggtagcg tctttcgccc
     2341 cactgtggcc tatctacatt ctcgtgcccg tggtctgtac ctgttcattt gacaggattc
     2401 ggggcagacc atacacaaca atcctgatgg gcatctactt tgtccttggg cttagctccg
     2461 ttggtatctt ttattgtctc atccacagac aagtgaaacg ggctgcccag gcccttgacc
     2521 agtacaaact gaggcaggct tctattcact ccaaccacgt ggcccgcaca gatgaggcaa
     2581 tgccaggccg atttcaggag ctcgactcac gccttgccag cggcgggcct tccgaaggaa
     2641 tctcttccga accagtatca gccgcgacaa cccagacgct tgaaggagac tcttctgagg
     2701 tcggtgacca gattaacagc aagcgagcca aacagatggc tgagaagagc ccgcctgaag
     2761 cctccgctaa ggcccagccc attaaaggcg ccagaagggc ccccgactcc tccagcgagt
     2821 ttggtaaagt gacacgcatg tgtttcgctg tctttctctg ctttgctctg agttatatcc
     2881 ccttcttgct tttgaatatt ctggacgcta gagtccaggc accgcgcgtt gttcacatgc
     2941 tggccgcaaa cctgacatgg ctgaacgggt gcatcaaccc cgtcttgtat gcagccatga
     3001 accggcaatt caggcaggcc tatgggtcta ttctgaagcg cggaccccgc tctttccatc
     3061 gcctgcacat cgataccggt ggacgcaccg gtgaattcga tgaggccagc accaccgtgt
     3121 ctaagaccga gacttctcag gtggcccctg cctaactcga gtctagaggg cccgtttaaa
     3181 cccgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc
     3241 ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg
     3301 aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg
     3361 acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta
     3421 tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta
     3481 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     3541 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     3601 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     3661 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     3721 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     3781 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     3841 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat
     3901 tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag
     3961 tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc
     4021 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct
     4081 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg
     4141 actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa
     4201 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat
     4261 atccattttc ggatctgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga
     4321 tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc
     4381 acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc
     4441 ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc
     4501 gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac
     4561 tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc
     4621 tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac
     4681 gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg
     4741 tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct
     4801 cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt
     4861 cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg
     4921 attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac
     4981 ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg
     5041 tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg
     5101 agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat
     5161 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     5221 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg
     5281 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     5341 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     5401 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct
     5461 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     5521 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     5581 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     5641 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     5701 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     5761 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     5821 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     5881 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     5941 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     6001 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     6061 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     6121 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     6181 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     6241 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac ggtatttggt
     6301 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     6361 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     6421 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     6481 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     6541 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     6601 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     6661 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     6721 ggccccag
//
