LOCUS       sequence-495869-        7350 bp    DNA     circular SYN 19-DEC-2025
DEFINITION  Expression vector for secreted human GPR124 GAIN domain, N-terminal 
            3xFLAG tag, C-terminal 6xHis tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7350)
  AUTHORS   Vallon M, Yuki K, Nguyen TD, Chang J, Yuan J, Siepe D, Miao Y, 
            Essler M, Noda M, Garcia KC, Kuo CJ
  TITLE     A RECK-WNT7 Receptor-Ligand Interaction Enables Isoform-Specific 
            Regulation of Wnt Bioavailability.
  JOURNAL   Cell Rep. 2018 Oct 9;25(2):339-349.e9. doi: 
            10.1016/j.celrep.2018.09.045.
  PUBMED    30304675
REFERENCE   2  (bases 1 to 7350)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 19, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.celrep.2018.09"; journalName: "Cell Rep"; date: 
            "2018-10-9- 9"; volume: "25"; issue: "2"; pages: "339-349.e9"
COMMENT     Sequence Label: p3xFLAG-CMV-9-GPR124 GAIN-His
FEATURES             Location/Qualifiers
     source          1..7350
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     366..385
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1148..1282
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1185..1204)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1239..1258
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1287..1306)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1288..1306)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1320..1336)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1320..1336)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1333..1355)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1344..1360
                     /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(1368..1374,1375..1392,1393..1398))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1413..1434
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1551..1568)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1722..2310)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1802..1821)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2481..3272,3273..3341))
                     /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     3104..3123
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3342..3446)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3560..3579)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3787..3808
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4071..4093
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4085..4102
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4086..4102
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        4263..4642
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4643..4846
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4797..4817
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             4877..4921
                     /codon_start=1
                     /product="leader sequence from mouse preprotrypsin"
                     /label=preprotrypsin leader
                     /translation="MSALLILALVGAAVA"
     CDS             4922..4987
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             6008..6025
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    6036..6658
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(6235..6254)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(6682..6702)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6700..6998
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             join(7052..7350,1..496)
                     /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(7106..7125)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat
       61 gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg
      121 catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga
      181 agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc gcatgcccga
      241 cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa
      301 tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga
      361 catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt
      421 cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct
      481 tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc gacgcccaac
      541 ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc
      601 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc
      661 gcccaccccg ggctcgatcc cctcgcgagt tggttcagct gctgcctgag gctggacgac
      721 ctcgcggagt tctaccggca gtgcaaatcc gtcggcatcc aggaaaccag cagcggctat
      781 ccgcgcatcc atgcccccga actgcaggag tggggaggca cgatggccgc tttggtcgac
      841 ccggacggga cgctcctgcg cctgatacag aacgaattgc ttgcaggcat ctcatgagtg
      901 tgtcttcccg ttttccgcct gaggtcactg cgtggatgga gcgctggcgc ctgctgcgcg
      961 acggcgagct gctcaccacc cactcgccaa gctggaaccg taaaaaggcc gcgttgctgg
     1021 cgtttttcca taggctccgc cgatcataat cagccatacc acatttgtag aggttttact
     1081 tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt
     1141 tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     1201 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     1261 tgtatcttat catgtctgga tcaattccct atagtgagtc gtattaaatt cgtaatcatg
     1321 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     1381 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     1441 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     1501 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     1561 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     1621 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     1681 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     1741 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     1801 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     1861 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     1921 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     1981 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     2041 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     2101 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     2161 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     2221 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     2281 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     2341 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     2401 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     2461 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     2521 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     2581 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     2641 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     2701 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     2761 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     2821 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     2881 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     2941 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     3001 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     3061 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     3121 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     3181 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     3241 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     3301 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     3361 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     3421 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgcgccc
     3481 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     3541 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     3601 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     3661 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
     3721 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaagag tccactatta
     3781 aagaacgtgg actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt
     3841 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     3901 aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttccatt cgccattcag
     3961 gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct tcgctattac gccagctggc
     4021 gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt cccagtcacg
     4081 acgttgtaaa acgacggcca gtgccaagct gatctataca ttgaatcaat attggcaatt
     4141 agccatatta gtcattggtt atatagcata aatcaatatt ggctattggc cattgcatac
     4201 gttgtatcta tatcataata tgtacattta tattggctca tgtccaatat gaccgccatg
     4261 ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag
     4321 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
     4381 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
     4441 gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca
     4501 tcaagtgtat catatgccaa gtccgccccc tattgacgtc aatgacggta aatggcccgc
     4561 ctggcattat gcccagtaca tgaccttacg ggactttcct acttggcagt acatctacgt
     4621 attagtcatc gctattacca tggtgatgcg gttttggcag tacaccaatg ggcgtggata
     4681 gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt
     4741 ttggcaccaa aatcaacggg actttccaaa atgtcgtaat aaccccgccc cgttgacgca
     4801 aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctcgtt tagtgaaccg
     4861 tcagaattaa ttcaccatgt ctgcacttct gatcctagct cttgttggag ctgcagttgc
     4921 tgactacaaa gaccatgacg gtgattataa agatcatgac atcgattaca aggatgacga
     4981 tgacaagctt gcggccgcga attcactgta caccttcgtg ctgatgccca tcaatgcctc
     5041 caatgcgctg accctggctc accagctgcg cgtgtacaca gccgaggccg ctagcttttc
     5101 agacatgatg gatgtagtct atgtggctca gatgatccag aaatttttgg gttatgtcga
     5161 ccagatcaaa gagctggtag aggtgatggt ggacatggcc agcaacctga tgctggtgga
     5221 cgagcacctg ctgtggctgg cccagcgcga ggacaaggcc tgcagccgca tcgtgggtgc
     5281 cctggagcgc attggggggg ccgccctcag cccccatgcc cagcacatct cagtgaatgc
     5341 gaggaacgtg gcattggagg cctacctcat caagccgcac agctacgtgg gcctgacctg
     5401 cacagccttc cagaggaggg agggaggggt gccgggcaca cggccaggaa gccctggcca
     5461 gaacccccca cctgagcccg agcccccagc tgaccagcag ctccgcttcc gctgcaccac
     5521 cgggaggccc aatgtttctc tgtcgtcctt ccacatcaag aacagcgtgg ccctggcctc
     5581 catccagctg cccccgagtc tattctcatc ccttccggct gccctggctc ccccggtgcc
     5641 cccagactgc accctgcaac tgctcgtctt ccgaaatggc cgcctcttcc acagccacag
     5701 caacacctcc cgccctggag ctgctgggcc tggcaagagg cgtggcgtgg ccacccccgt
     5761 catcttcgca ggaaccagtg gctgtggcgt gggaaacctg acagagccag tggccgtttc
     5821 gctgcggcac tgggctgagg gagccgaacc tgtggccgct tggtggagcc aggaggggcc
     5881 cggggaggct gggggctgga cctcggaggg ctgccagctc cgctccagcc agcccaatgt
     5941 cagcgccctg cactgccagc acttgggcaa tgtggccgtg ctcatggagc tggatggggg
     6001 gggttctcat catcatcatc atcattaagg atcccgggtg gcatccctgt gacccctccc
     6061 cagtgcctct cctggccctg gaagttgcca ctccagtgcc caccagcctt gtcctaataa
     6121 aattaagttg catcattttg tctgactagg tgtccttcta taatattatg gggtggaggg
     6181 gggtggtatg gagcaagggg caagttggga agacaacctg tagggcctgc ggggtctatt
     6241 gggaaccaag ctggagtgca gtggcacaat cttggctcac tgcaatctcc gcctcctggg
     6301 ttcaagcgat tctcctgcct cagcctcccg agttgttggg attccaggca tgcatgacca
     6361 ggctcagcta atttttgttt ttttggtaga gacggggttt caccatattg gccaggctgg
     6421 tctccaactc ctaatctcag gtgatctacc caccttggcc tcccaaattg ctgggattac
     6481 aggcgtgaac cactgctccc ttccctgtcc ttctgatttt aaaataacta taccagcagg
     6541 aggacgtcca gacacagcat aggctacctg gccatgccca accggtggga catttgagtt
     6601 gcttgcttgg cactgtcctc tcatgcgttg ggtccactca gtagatgcct gttgaattgg
     6661 gtacgcggcc agcttggctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     6721 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
     6781 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     6841 ccatagtccc gcccctaact ccgcccatcc cgcctccgct aactccgccc agttccgccc
     6901 attctccgcc ccatggctga ctaatttttt ttatttatgc aggccgaggc cgcctcggcc
     6961 tctgagctat tccagaagta gtgaggaggc ttttttggag gaattgatca gcttgggatc
     7021 tgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg
     7081 ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg
     7141 ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa
     7201 gaccgacctg tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct
     7261 ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga
     7321 ctggctgcta ttgggcgaag tgccggggca
//
