LOCUS       sequence-430747-        6971 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 6971)
  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 6971)
  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: ADRB1-DuET
FEATURES             Location/Qualifiers
     source          1..6971
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     405..424
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(643..747)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(822..841)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1013..1392
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1393..1596
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1547..1567
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1641..1660
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1641..1659
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3340..3357)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3346..3570
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3616..4044
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3703..3722)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3913..3934
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4053..4073)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4058..4387
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4238..4373
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4300..4319
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4454..5248
                     /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(4508..4527)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5118..5137
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5422..5543
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5459..5478)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5513..5532
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5592..5608)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5592..5608)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5605..5627)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5616..5632
                     /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(5640..5646,5647..5664,5665..5670))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5685..5706
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5823..5840)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5994..6582)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6074..6093)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6753..6971,1..573,574..642))
                     /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 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
       61 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
      121 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
      181 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
      241 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
      301 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
      361 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
      421 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
      481 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
      541 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
      601 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
      661 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
      721 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcga
      781 cggatcggga gatctcccga tcccctatgg tgcactctca gtacaatctg ctctgatgcc
      841 gcatagttaa gccagtatct gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg
      901 agcaaaattt aagctacaac aaggcaaggc ttgaccgaca tttgcatgaa gaatctgctt
      961 agggttaggc gttttgcgct gcttcgcgat gtacgggcca gatatacgcg ttgacattga
     1021 ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg
     1081 gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc
     1141 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat
     1201 tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat
     1261 catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
     1321 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc
     1381 gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac
     1441 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa
     1501 aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt
     1561 aggcgtgtac ggtgggaggt ctatataagc agagctctct ggctaactag agaacccact
     1621 gcttactggc ttatcgaaat taatacgact cactataggg agacccaagc tggctagcgt
     1681 ttaaacttaa gcttagcgcc accatgaaga cgatcatcgc cctgagctac atcttctgcc
     1741 tggtattcgc cgactacaag gacgatgatg acgccagcat cgatatgggg gcaggagtac
     1801 ttgtgctggg ggccagcgag ccgggaaatc ttagctctgc tgcacctctg cctgatggag
     1861 ctgctacagc cgcaagactg ctggttccgg ctagcccccc tgcgtcattg ctcccgcccg
     1921 ccagtgagag cccagagccc ctgagccagc aatggaccgc tggcatggga ctgctcatgg
     1981 ccttgattgt gctgcttata gtagcaggaa atgtgttggt gatagttgct atcgccaaaa
     2041 ccccccgcct tcaaaccctc acaaatcttt tcataatgag cctggcttcc gccgacctgg
     2101 tgatgggact gctcgtggtt ccattcggcg ctacaatcgt ggtttggggg cgatgggagt
     2161 atggatcctt cttctgcgag ctctggacta gtgtagatgt gttgtgcgtt accgcatcaa
     2221 tagagactct gtgcgtaatc gccctggatc gctaccttgc aattaccagt ccgttcagat
     2281 atcaaagcct gctgacaaga gcccgggcca gaggactggt ctgtaccgtg tgggccatca
     2341 gcgccctggt gagcttcctc ccgatcctca tgcattggtg gcgagcagag tctgatgagg
     2401 ctagaaggtg ttacaatgac cctaagtgct gtgacttcgt caccaatcgc gcttacgcga
     2461 tcgcgtccag tgtcgtatct ttctacgtgc ctttgtgcat catggccttc gtttatctcc
     2521 gcgttttccg ggaggcacag aaacaggtca aaaagattga ttcttgtgaa cggaggtttc
     2581 tgggtggtcc cgccagaccc ccatccccgt cacccagccc cgtacccgct cccgcccccc
     2641 cacccggacc acctagaccc gcagcagctg ccgctaccgc accgctggcc aatggtcggg
     2701 ctggaaaaag aagaccaagc cgcctcgttg ccctgcggga acagaaagcg cttaaaacct
     2761 tgggcattat catgggagtc tttacattgt gctggctccc tttcttcctg gccaatgttg
     2821 ttaaggcatt ccacagagaa ctcgtgccgg atagattgtt cgtcttcttt aactggctgg
     2881 gatacgccaa ctctgctttc aatcccatta tctactgccg ctctcccgat ttcagaaagg
     2941 ccttccaagg cctcctttgt tgcgcccgcc gggccgctag aaggaggcac gcaactcacg
     3001 gggaccggcc aagggcgtcc ggatgtctcg cacgcccggg cccaccaccc tccccaggag
     3061 ctgcctccga cgatgacgac gacgatgtgg tcggagctac accaccagcg cgcctgctgg
     3121 agccctgggc aggttgcaat ggcggcgctg ctgcagattc tgacagctcc ttggacgaac
     3181 cttgtagacc cggctttgct tcagagtcca aggtcatcga taccggtgga cgcaccggtg
     3241 aattcgatga ggccagcacc accgtgtcta agaccgagac ttctcaggtg gcccctgcct
     3301 aactcgagtc tagagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt
     3361 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     3421 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     3481 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
     3541 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct
     3601 ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     3661 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     3721 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     3781 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     3841 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     3901 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
     3961 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     4021 tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa
     4081 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     4141 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     4201 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     4261 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     4321 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     4381 ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcaag agacaggatg
     4441 aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt
     4501 ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt
     4561 gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc
     4621 cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc
     4681 ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga
     4741 agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat
     4801 ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca
     4861 agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga
     4921 tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc
     4981 gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat
     5041 catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga
     5101 ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg
     5161 ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt
     5221 ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa
     5281 gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg
     5341 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     5401 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
     5461 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     5521 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
     5581 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     5641 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     5701 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     5761 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     5821 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     5881 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     5941 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     6001 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     6061 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     6121 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     6181 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     6241 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     6301 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     6361 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     6421 ggctacacta gaagaacggt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     6481 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     6541 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     6601 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     6661 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     6721 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     6781 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     6841 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     6901 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     6961 agtggtcctg c
//
