LOCUS       sequence-430831-        6626 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 6626)
  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 6626)
  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: GPR6-DuET
FEATURES             Location/Qualifiers
     source          1..6626
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     449..468
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(687..791)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(866..885)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1057..1436
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1437..1640
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1591..1611
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1685..1704
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1685..1703
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3039..3056)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3045..3269
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3315..3743
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3402..3421)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3612..3633
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3752..3772)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3757..4086
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3937..4072
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3999..4018
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4153..4947
                     /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(4207..4226)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4817..4836
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5121..5242
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5158..5177)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5212..5231
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5291..5307)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5291..5307)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5304..5326)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5315..5331
                     /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(5339..5345,5346..5363,5364..5369))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5384..5405
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5522..5539)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5693..6281)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5773..5792)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6452..6626,1..617,618..686))
                     /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 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
       61 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
      121 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
      181 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
      241 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
      301 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
      361 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
      421 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
      481 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
      541 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
      601 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
      661 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
      721 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
      781 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc gggagatctc
      841 ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt
      901 atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta
      961 caacaaggca aggcttgacc gacatttgca tgaagaatct gcttagggtt aggcgttttg
     1021 cgctgcttcg cgatgtacgg gccagatata cgcgttgaca ttgattattg actagttatt
     1081 aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat
     1141 aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa
     1201 taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg
     1261 agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc
     1321 cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct
     1381 tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga
     1441 tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa
     1501 gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc
     1561 caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg
     1621 aggtctatat aagcagagct ctctggctaa ctagagaacc cactgcttac tggcttatcg
     1681 aaattaatac gactcactat agggagaccc aagctggcta gcgtttaaac ttaagcttag
     1741 cgccaccatg aagacgatca tcgccctgag ctacatcttc tgcctggtat tcgccgacta
     1801 caaggacgat gatgacgcca gcatcgatat gaatgctagc gctgcatccc tgaatgactc
     1861 ccaggtagtg gtggtagcag ctgaaggggc agctgcagca gccaccgctg ccgggggccc
     1921 tgacacagga gagtgggggc cccctgccgc ggcggctctt ggtgccggcg gaggggccaa
     1981 cggcagtctc gaactcagta gtcagctgtc tgccggaccc cccggtctct tgttgcctgc
     2041 cgtaaaccct tgggatgtcc ttctgtgcgt gagtggaacg gttatcgctg gagagaatgc
     2101 cctcgtggtg gccctgatcg catccactcc cgctctccgc accccaatgt tcgttctggt
     2161 tggatcactg gcgacagccg atcttctcgc cggctgcggg ctcatcctcc acttcgtgtt
     2221 ccaatatttg gtaccgtccg aaaccgtatc actcttgacc gtcgggttcc ttgtcgcaag
     2281 ctttgccgcc tcagtcagtt cccttttggc catcactgtg gacagatacc tgtcattgta
     2341 taacgcactt acctattaca gcagaaggac attgctgggc gtccacctgc tcctggccgc
     2401 tacctggacc gtgagtctgg ggctcggact ccttcctgtc ctcgggtgga actgcctggc
     2461 agaaagggct gcctgttctg ttgtgaggcc cctggcacgc tctcatgtgg cactgctgag
     2521 tgccgccttc ttcatggtct ttgggatcat gctgcacctc tacgttcgca tttgccaggt
     2581 ggtgtggagg catgctcatc agatagcctt gcaacagcac tgcctcgcgc cacctcatct
     2641 ggcggctact cgcaaagggg tcggtaccct ggcagtggta ctgggcacct ttggggctag
     2701 ttggctgcct ttcgccatct actgtgtcgt gggaagccat gaggatcccg ccgtctatac
     2761 atacgcgacg ctcctgccag ccacatataa cagcatgatc aaccctataa tatatgcatt
     2821 ccggaatcag gagatccaga gagccctgtg gctgctgttg tgcggctgtt ttcagtctaa
     2881 agtgcctttc cggtcccgaa gccccagtga ggtcatcgat accggtggac gcaccggtga
     2941 attcgatgag gccagcacca ccgtgtctaa gaccgagact tctcaggtgg cccctgccta
     3001 actcgagtct agagggcccg tttaaacccg ctgatcagcc tcgactgtgc cttctagttg
     3061 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     3121 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     3181 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag
     3241 gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc
     3301 tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     3361 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     3421 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     3481 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     3541 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     3601 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     3661 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
     3721 ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag
     3781 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     3841 aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     3901 tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
     3961 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc
     4021 gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt
     4081 tgcaaaaagc tcccgggagc ttgtatatcc attttcggat ctgatcaaga gacaggatga
     4141 ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg
     4201 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg
     4261 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc
     4321 ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct
     4381 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa
     4441 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg
     4501 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa
     4561 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat
     4621 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg
     4681 cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc
     4741 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac
     4801 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg
     4861 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc
     4921 tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag
     4981 cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg
     5041 gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc
     5101 tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca
     5161 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     5221 ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg
     5281 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     5341 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     5401 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     5461 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     5521 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     5581 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     5641 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     5701 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     5761 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     5821 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     5881 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     5941 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     6001 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     6061 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     6121 gctacactag aagaacggta tttggtatct gcgctctgct gaagccagtt accttcggaa
     6181 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     6241 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     6301 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     6361 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     6421 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     6481 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     6541 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     6601 gctcaccggc tccagattta tcagca
//
