LOCUS       sequence-495881-        8597 bp    DNA     circular SYN 19-DEC-2025
DEFINITION  Expression vector for secreted human GPR124 ECD D364E (RGD -> RGE), 
            N-terminal 3xFLAG tag, C-terminal 6xHis tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8597)
  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 8597)
  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-sGPR124 D364E-His
FEATURES             Location/Qualifiers
     source          1..8597
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     28..47
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        28..46
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    52..186
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(76..95)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     130..149
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(838..1632)
                     /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(949..968)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1559..1578
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(1686..1984)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     primer_bind     1982..2002
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    2026..2648
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     2430..2449
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     CDS             complement(2760..2771)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             complement(4944..5009)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             complement(5010..5054)
                     /codon_start=1
                     /product="leader sequence from mouse preprotrypsin"
                     /label=preprotrypsin leader
                     /translation="MSALLILALVGAAVA"
     promoter        complement(5085..5288)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(5114..5134)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        5289..5668
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     complement(5829..5846)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5829..5845)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5838..5860)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(6123..6144)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6352..6371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        6485..6589
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6590..6658,6659..7450)
                     /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     complement(6808..6827)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7621..8209
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8110..8129
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8363..8380
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    8497..8518
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(8533..8538,8539..8556,8557..8563)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8571..8587
                     /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)."
     primer_bind     join(8576..8597,1)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     join(8595..8597,1..14)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     join(8595..8597,1..14)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 gaaacagcta tgacatgatt acgaatttaa tacgactcac tatagggaat tgatccagac
       61 atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc
      121 tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa
      181 caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga ggtgtgggag
      241 gttttttaaa gcaagtaaaa cctctacaaa tgtggtatgg ctgattatga tcggcggagc
      301 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc agcttggcga gtgggtggtg
      361 agcagctcgc cgtcgcgcag caggcgccag cgctccatcc acgcagtgac ctcaggcgga
      421 aaacgggaag acacactcat gagatgcctg caagcaattc gttctgtatc aggcgcagga
      481 gcgtcccgtc cgggtcgacc aaagcggcca tcgtgcctcc ccactcctgc agttcggggg
      541 catggatgcg cggatagccg ctgctggttt cctggatgcc gacggatttg cactgccggt
      601 agaactccgc gaggtcgtcc agcctcaggc agcagctgaa ccaactcgcg aggggatcga
      661 gcccggggtg ggcgaagaac tccagcatga gatccccgcg ctggaggatc atccagccgg
      721 cgtcccggaa aacgattccg aagcccaacc tttcatagaa ggcggcggtg gaatcgaaat
      781 ctcgtgatgg caggttgggc gtcgcttggt cggtcatttc gaaccccaga gtcccgctca
      841 gaagaactcg tcaagaaggc gatagaaggc gatgcgctgc gaatcgggag cggcgatacc
      901 gtaaagcacg aggaagcggt cagcccattc gccgccaagc tcttcagcaa tatcacgggt
      961 agccaacgct atgtcctgat agcggtccgc cacacccagc cggccacagt cgatgaatcc
     1021 agaaaagcgg ccattttcca ccatgatatt cggcaagcag gcatcgccat gggtcacgac
     1081 gagatcctcg ccgtcgggca tgcgcgcctt gagcctggcg aacagttcgg ctggcgcgag
     1141 cccctgatgc tcttcgtcca gatcatcctg atcgacaaga ccggcttcca tccgagtacg
     1201 tgctcgctcg atgcgatgtt tcgcttggtg gtcgaatggg caggtagccg gatcaagcgt
     1261 atgcagccgc cgcattgcat cagccatgat ggatactttc tcggcaggag caaggtgaga
     1321 tgacaggaga tcctgccccg gcacttcgcc caatagcagc cagtcccttc ccgcttcagt
     1381 gacaacgtcg agcacagctg cgcaaggaac gcccgtcgtg gccagccacg atagccgcgc
     1441 tgcctcgtcc tgcagttcat tcagggcacc ggacaggtcg gtcttgacaa aaagaaccgg
     1501 gcgcccctgc gctgacagcc ggaacacggc ggcatcagag cagccgattg tctgttgtgc
     1561 ccagtcatag ccgaatagcc tctccaccca agcggccgga gaacctgcgt gcaatccatc
     1621 ttgttcaatc atgcgaaacg atcctcatcc tgtctcttga tcagatccca agctgatcaa
     1681 ttcctccaaa aaagcctcct cactacttct ggaatagctc agaggccgag gcggcctcgg
     1741 cctgcataaa taaaaaaaat tagtcagcca tggggcggag aatgggcgga actgggcgga
     1801 gttagcggag gcgggatggg cggagttagg ggcgggacta tggttgctga ctaattgaga
     1861 tgcatgcttt gcatacttct gcctgctggg gagcctgggg actttccaca cctggttgct
     1921 gactaattga gatgcatgct ttgcatactt ctgcctgctg gggagcctgg ggactttcca
     1981 caccctaact gacacacatt ccacagccaa gctggccgcg tacccaattc aacaggcatc
     2041 tactgagtgg acccaacgca tgagaggaca gtgccaagca agcaactcaa atgtcccacc
     2101 ggttgggcat ggccaggtag cctatgctgt gtctggacgt cctcctgctg gtatagttat
     2161 tttaaaatca gaaggacagg gaagggagca gtggttcacg cctgtaatcc cagcaatttg
     2221 ggaggccaag gtgggtagat cacctgagat taggagttgg agaccagcct ggccaatatg
     2281 gtgaaacccc gtctctacca aaaaaacaaa aattagctga gcctggtcat gcatgcctgg
     2341 aatcccaaca actcgggagg ctgaggcagg agaatcgctt gaacccagga ggcggagatt
     2401 gcagtgagcc aagattgtgc cactgcactc cagcttggtt cccaatagac cccgcaggcc
     2461 ctacaggttg tcttcccaac ttgccccttg ctccatacca cccccctcca ccccataata
     2521 ttatagaagg acacctagtc agacaaaatg atgcaactta attttattag gacaaggctg
     2581 gtgggcactg gagtggcaac ttccagggcc aggagaggca ctggggaggg gtcacaggga
     2641 tgccacccgg gatcctctag agtcgactgg taccgatatc agatctatcg atgaattcgc
     2701 ggccgcaagc ttggtaccga gctcggatcc actagtaacg gccgccagtg tgctggaatt
     2761 cgcccttcta tcggccgtta cagctccata agcacggcca cattgcccaa gtgctggcag
     2821 tacagggagc tgacgttggg ctggctgtag cggagcctgc agccttcaga actccagcct
     2881 ccaggcccgt cctggttcca ccaagctgcc atggggtcag ctccttcagc ccagtgcctc
     2941 agtgacacag ccacgggctc cgtcaagttt cccacaccac agccactggt tcctgcaaat
     3001 atgactgggg tggccacacc acgcctcttg ccaggcccag ctgctccagg acgggaagtg
     3061 ttgttgccgt ggctgcggaa aagacggcca tttctgaaga ccagcagttg cagggtgcaa
     3121 tctggaggga ctgggggagc cagggcagcc ggaagggttg agaacaggct ggggggcagc
     3181 tggatggagg ccagggccac gctattcttg atgtggaagg atgacagaga aatgttgggc
     3241 ctcccagtgg tgcagcggaa cctaagctgc tgatcagcta ggggttcggg ctccactggg
     3301 gcatcctggc cgacgctgct tggctgtgca cccgacactc ctacctccct cctctggaaa
     3361 gccgtgcaag tcagacccac gtagctgtga ggcttgatga ggtaggcctc cagtgccaca
     3421 tttcttgaat tcacagagat gtgctgggca tgggggctaa gagcagctcc tccgattcgc
     3481 tccagggcac ccacaatgcc actgcaggct ttgtcttctc tctgggccag ccacagaagg
     3541 tgctcatcca ccagcatcag gttgctggcc atgtccacca tcacctccac cagctccttg
     3601 atctggtcaa cgtaacccaa aaacttctgg atcatctgag ccacatagac cacgtccatc
     3661 atgtctgaga agctggcggc ctctgcggta tacactcgca gctggtgggc caacgtcaat
     3721 gcattggagg cgttgatggg catcagcaca aaggtgtaga gcacccgagt gatgtcattg
     3781 gtgtacagac agtgcgagta gtcccctggc tcccagcggc cagctcggtc acacctgcgt
     3841 gaggctcggg tacccggggc tcccccactc aagggcacag aggtgaaggg gtactgtaaa
     3901 caggactggt aagccgtgat gccagccaag gttcggggcc acctgaagtc cccacggttg
     3961 ttggtcaccc gctctgcagg gcagtaggag gcagaggtct ccagcactac tatctccacc
     4021 ttcttgctgg tgttgccttg gactgtggac acggagcact cccattcacc agaggcccac
     4081 acaccaatgt gagacagggt cagctcactg gtgatgaagg tgcaatcatg gataaggttt
     4141 tcagcaagga cgatgccggc ctgctcatcg ctctccatag gagccccatt gtggtaccag
     4201 tggatccggg tatcgttgcc caagtagctg gctgagcact ggaagggcag gcggtcaccc
     4261 tggaacacca cttggcgaag ggatgggatg aggtagtggg tgtgcagttc cagggcccct
     4321 tcacagcgaa gctgggactc ctggaggctg ctcagggcat gggcgtgcag ggcactgggg
     4381 taggcacaga gtgtgcgctc agacagctgc agggagtgat tccgggccca gggcagcagc
     4441 cagcgcaggc ggcagtcaca ggtcagaaac tcagtaccaa agtccacaat cttaagggct
     4501 ggcagctcat caaagacccc aggttgcaga ctagagtaga tgtttccaga tatgtttagt
     4561 ctgagaagtc tagggagccc ttgaaatgtc tcagaggtga gacagccaat ccgattgttg
     4621 gagagatcta agcgttttag ctctcccaga cctaggaagg ctccaggctg cacagtgctg
     4681 atgacattgc tcctcaggtc caacttctcc aacagggaca gtcccaagaa ggatccattg
     4741 cggagcccag taatcttgtt gttgctcaag agcaaggtga tggtgccgtt tggcagaagg
     4801 ccgggatctg gaggttcggg gagatcccca ctgccgcaca ccaccctcct tcgagccggg
     4861 ttgtgggcgc cgccacttag tcccttgggc cgctccccag agcacttgca accgcggata
     4921 gggaccgggc agccaggcgc aagcttgtca tcgtcatcct tgtaatcgat gtcatgatct
     4981 ttataatcac cgtcatggtc tttgtagtca gcaactgcag ctccaacaag agctaggatc
     5041 agaagtgcag acatggtgaa ttaattctga cggttcacta aacgagctct gcttatatag
     5101 acctcccacc gtacacgcct accgcccatt tgcgtcaacg gggcggggtt attacgacat
     5161 tttggaaagt cccgttgatt ttggtgccaa aacaaactcc cattgacgtc aatggggtgg
     5221 agacttggaa atccccgtga gtcaaaccgc tatccacgcc cattggtgta ctgccaaaac
     5281 cgcatcacca tggtaatagc gatgactaat acgtagatgt actgccaagt aggaaagtcc
     5341 cgtaaggtca tgtactgggc ataatgccag gcgggccatt taccgtcatt gacgtcaata
     5401 gggggcggac ttggcatatg atacacttga tgtactgcca agtgggcagt ttaccgtaaa
     5461 tactccaccc attgacgtca atggaaagtc cctattggcg ttactatggg aacatacgtc
     5521 attattgacg tcaatgggcg ggggtcgttg ggcggtcagc caggcgggcc atttaccgta
     5581 agttatgtaa cgcggaactc catatatggg ctatgaacta atgaccccgt aattgattac
     5641 tattaataac tagtcaataa tcaatgtcaa catggcggtc atattggaca tgagccaata
     5701 taaatgtaca tattatgata tagatacaac gtatgcaatg gccaatagcc aatattgatt
     5761 tatgctatat aaccaatgac taatatggct aattgccaat attgattcaa tgtatagatc
     5821 agcttggcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa
     5881 cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc
     5941 accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggaaatt gtaagcgtta
     6001 atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg
     6061 ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg
     6121 ttccagtttg gaacaagagt ccacgttctt taatagtgga ctcttaaaga acgtggactc
     6181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
     6241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
     6301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
     6361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
     6421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa
     6481 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     6541 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     6601 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     6661 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     6721 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     6781 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     6841 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     6901 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     6961 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     7021 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     7081 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     7141 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     7201 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     7261 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     7321 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     7381 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     7441 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     7501 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     7561 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     7621 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     7681 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     7741 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     7801 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     7861 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     7921 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     7981 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     8041 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     8101 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     8161 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     8221 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     8281 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     8341 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
     8401 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
     8461 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
     8521 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
     8581 ataacaattt cacacag
//
