LOCUS       sequence-430799-        7625 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 7625)
  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 7625)
  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: FSHR-DuET
FEATURES             Location/Qualifiers
     source          1..7625
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     181..200
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(419..523)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(598..617)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        789..1168
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1169..1372
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1323..1343
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1417..1436
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1417..1435
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3770..3787)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3776..4000
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4046..4474
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4133..4152)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4343..4364
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4483..4503)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4488..4817
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4668..4803
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4730..4749
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4884..5678
                     /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(4938..4957)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5548..5567
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5852..5973
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5889..5908)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5943..5962
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6022..6038)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6022..6038)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6035..6057)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6046..6062
                     /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(6070..6076,6077..6094,6095..6100))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6115..6136
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6253..6270)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6424..7012)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6504..6523)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7183..7625,1..349,350..418))
                     /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 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
       61 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
      121 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
      181 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
      241 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
      301 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
      361 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
      421 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
      481 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
      541 tgccacctga cgtcgacgga tcgggagatc tcccgatccc ctatggtgca ctctcagtac
      601 aatctgctct gatgccgcat agttaagcca gtatctgctc cctgcttgtg tgttggaggt
      661 cgctgagtag tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacatttg
      721 catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata
      781 tacgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
      841 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
      901 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
      961 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
     1021 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
     1081 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
     1141 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg
     1201 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
     1261 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt
     1321 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctctctggct
     1381 aactagagaa cccactgctt actggcttat cgaaattaat acgactcact atagggagac
     1441 ccaagctggc tagcgtttaa acttaagctt agcgccacca tgaagacgat catcgccctg
     1501 agctacatct tctgcctggt attcgccgac tacaaggacg atgatgacgc cagcatcgat
     1561 atggcgctgc tgcttgttag tctccttgcc tttttgagcc tgggatccgg gtgtcaccat
     1621 aggatctgtc attgttcaaa ccgggtgttc ttgtgtcaag agtccaaagt caccgaaatc
     1681 ccatctgatc tgccaagaaa tgccattgag cttagattcg tgctgactaa actcagggtg
     1741 attcagaagg gcgccttctc tgggttcgga gacctggaaa aaatcgagat ctcccagaat
     1801 gacgtgctgg aagtaatcga ggctgatgtg tttagcaacc tgccaaaact gcacgaaatc
     1861 aggattgaga aagcgaacaa tcttctgtat ataaatcccg aggcattcca gaatttgcct
     1921 aacctccaat atctcctgat atccaatacc ggcatcaagc acctccccga tgtgcataag
     1981 atacactctt tgcagaaggt cctgctggac atccaggata atattaacat ccatacaatc
     2041 gagcgaaact ctttcgtggg ccttagcttt gaatccgtga ttttgtggct gaacaagaat
     2101 ggcattcaag agatccacaa ttgcgccttt aacgggaccc agctggacga gctcaatctg
     2161 agcgacaata acaatctgga ggagctcccc aacgacgtgt ttcatggcgc gagtggaccc
     2221 gttatcctcg acattagccg gacacgcatc cattctcttc catcctacgg actggaaaac
     2281 cttaagaagc tcagggcgcg atccacctat aacctgaaaa agctgccgac acttgagaag
     2341 ttggtcgccc tcatggaggc tagcctcacc tatccatccc attgttgtgc cttcgctaat
     2401 tggaggcggc aaatctctga acttcaccca atctgcaata aatccatcct caggcaagag
     2461 gtggattaca tgacccaggc cagaggtcag cggtcaagcc tggcagaaga caatgagtcc
     2521 tcttacagcc gcgggttcga tatgacctat acagagtttg actacgacct gtgcaacgag
     2581 gtggtggatg tcacgtgttc tccaaagccc gacgccttta acccatgtga ggacatcatg
     2641 ggctacaaca tcctgagagt gctgatatgg tttatctcaa tcctggctat taccggcaat
     2701 atcatcgtcc tggtcattct tactacatcc caatataagc tcacggtgcc acgctttctg
     2761 atgtgcaacc tcgcttttgc cgacctttgt atcgggatct atctcctgct gatcgcatcc
     2821 gttgacatcc acacaaagag ccagtaccac aattacgcga ttgattggca gaccggggct
     2881 ggctgtgacg ctgcaggatt ctttaccgtg tttgcgtcag agttgagtgt gtacaccctg
     2941 acagcgatta ccctggagcg gtggcacacc attacccacg cgatgcaact ggattgcaag
     3001 gtccaactgc ggcatgctgc ttcagtgatg gttatgggtt ggattttcgc tttcgcagca
     3061 gcactgtttc caatcttcgg catcagttct tacatgaaag tttccatctg tctgccaatg
     3121 gacattgaca gcccccttag ccagctgtac gtcatgtccc ttctggtgct caacgtgctg
     3181 gccttcgtgg tcatttgtgg atgctacatc catatttacc tgactgttcg gaatcccaac
     3241 atcgtgtctt ccagctccga cactaggata gccaagagga tggctatgct gatcttcacg
     3301 gatttcctct gcatggcacc gatttcattc ttcgccatct ctgcctccct gaaggtgccg
     3361 ctgattacgg tgtctaaagc taagattctg cttgtattgt tccacccaat taactcttgt
     3421 gccaatccat tcctgtacgc aatatttacc aaaaacttca ggcgagattt ttttatcctc
     3481 ctgtcaaagt gtgggtgcta tgagatgcaa gctcaaattt atagaactga aacttccagt
     3541 acagtacaca atacgcaccc tcggaacggc cattgctcct cagctccgcg ggtaacaagc
     3601 ggatccacat acattctggt cccactgtca cacctggccc agaacatcga taccggtgga
     3661 cgcaccggtg aattcgatga ggccagcacc accgtgtcta agaccgagac ttctcaggtg
     3721 gcccctgcct aactcgagtc tagagggccc gtttaaaccc gctgatcagc ctcgactgtg
     3781 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     3841 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt
     3901 aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa
     3961 gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc
     4021 agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt
     4081 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     4141 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     4201 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     4261 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     4321 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     4381 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     4441 aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag
     4501 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     4561 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     4621 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     4681 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     4741 aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     4801 gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcaag
     4861 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     4921 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     4981 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     5041 tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga
     5101 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     5161 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     5221 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     5281 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     5341 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     5401 ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     5461 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     5521 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     5581 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     5641 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat
     5701 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta
     5761 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     5821 ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta
     5881 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     5941 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag
     6001 ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     6061 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     6121 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     6181 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     6241 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     6301 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     6361 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     6421 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     6481 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     6541 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     6601 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     6661 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     6721 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     6781 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     6841 gcctaactac ggctacacta gaagaacggt atttggtatc tgcgctctgc tgaagccagt
     6901 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     6961 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     7021 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     7081 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     7141 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     7201 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     7261 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     7321 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     7381 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     7441 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     7501 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     7561 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     7621 tccga
//
