LOCUS       sequence-360374-        6957 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses FLAG tagged FZD4 swapped with FZD3 CRDlinker domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6957)
  AUTHORS   Ko SB, Mihara E, Park Y, Roh K, Kang C, Takagi J, Bang I, Choi HJ
  TITLE     Functional role of the Frizzled linker domain in the Wnt signaling 
            pathway.
  JOURNAL   Commun Biol. 2022 May 5;5(1):421. doi: 10.1038/s42003-022-03370-4.
  PUBMED    35513706
REFERENCE   2  (bases 1 to 6957)
  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: "Commun 
            Biol."; date: "2022-05-5"; volume: "5(1):421. doi"; pages: " 
            10.1038/s42003-022-03370-4"
COMMENT     Sequence Label: pcDNA FZD4_3CRDlinker
FEATURES             Location/Qualifiers
     source          1..6957
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             394..411
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(434..451)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    440..664
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      710..1138
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(797..816)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1007..1028
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1147..1167)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1152..1481
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1332..1467
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1394..1413
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             1548..2342
                     /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(1602..1621)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2212..2231
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    2518..2639
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2555..2574)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2609..2628
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2688..2704)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2688..2704)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2701..2723)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2712..2728
                     /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(2736..2742,2743..2760,2761..2766))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2781..2802
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2919..2936)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3090..3678)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3170..3189)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3849..4640,4641..4709))
                     /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     4472..4491
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4710..4814)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4889..4908)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        5080..5459
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5460..5663
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5614..5634
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5708..5727
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5708..5726
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5761..5805
                     /codon_start=1
                     /product="leader sequence from mouse preprotrypsin"
                     /label=preprotrypsin leader
                     /translation="MSALLILALVGAAVA"
     CDS             5806..5829
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
ORIGIN
        1 ttggtggcct tattcaaaat tcggtccaat cttcaaaaag acgggacaaa gacagacaag
       61 ttggaaaggc taatggtcaa gatcggggtc ttctcagtac tgtacacggt tcctgcaacc
      121 tgtgtgattg cctgttattt ctatgaaatc tcaaactggg cactctttcg atattctgca
      181 gatgactcaa acatggcagt tgaaatgttg aaaattttta tgtctttgct cgtgggcatc
      241 acttcaggca tgtggatttg gtctgccaaa actcttcaca cgtggcaaaa gtgttctaac
      301 cgattggtga attctgggaa ggtaaagaga gagaagaggg ggaatggttg ggtgaagcca
      361 ggaaaaggca acgagactgt ggtatagctc gagcatcatc accatcacca ttgagtttaa
      421 acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
      481 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
      541 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag
      601 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct
      661 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt
      721 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
      781 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
      841 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
      901 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
      961 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     1021 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     1081 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa
     1141 ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     1201 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     1261 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     1321 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     1381 gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga
     1441 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta
     1501 tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag
     1561 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
     1621 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
     1681 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag
     1741 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
     1801 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
     1861 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
     1921 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
     1981 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
     2041 tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg
     2101 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
     2161 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
     2221 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
     2281 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
     2341 gagcgggact ctggggttcg cgaaatgacc gaccaagcga cgcccaacct gccatcacga
     2401 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
     2461 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac
     2521 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     2581 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     2641 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt
     2701 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     2761 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     2821 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     2881 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     2941 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     3001 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     3061 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     3121 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     3181 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     3241 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     3301 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     3361 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     3421 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     3481 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     3541 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     3601 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     3661 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     3721 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     3781 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     3841 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     3901 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     3961 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     4021 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     4081 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     4141 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     4201 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     4261 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     4321 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     4381 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     4441 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     4501 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     4561 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     4621 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     4681 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     4741 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     4801 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtcgacgg atcgggagat
     4861 ctcccgatcc cctatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     4921 agtatctgct ccctgcttgt gtgttggagg tcgctgagta gtgcgcgagc aaaatttaag
     4981 ctacaacaag gcaaggcttg accgacaatt gcatgaagaa tctgcttagg gttaggcgtt
     5041 ttgcgctgct tcgcgatgta cgggccagat atacgcgttg acattgatta ttgactagtt
     5101 attaatagta atcaattacg gggtcattag ttcatagccc atatatggag ttccgcgtta
     5161 cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt
     5221 caataatgac gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg
     5281 tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagta
     5341 cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga
     5401 ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgg
     5461 tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca cggggatttc
     5521 caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact
     5581 ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg cgtgtacggt
     5641 gggaggtcta tataagcaga gctctctggc taactagaga acccactgct tactggctta
     5701 tcgaaattaa tacgactcac tatagggaga cccaagctgg ctagttaagc ttggtaccga
     5761 atgtctgcac ttctgatcct agctcttgtt ggagctgcag ttgctgacta caaagacgat
     5821 gacgacaagc ttgcggccgc acacagtttg ttttcttgtg aacctattac cttgaggatg
     5881 tgccaagatt tgccttataa tactaccttc atgcctaatc ttctgaatca ttatgaccaa
     5941 cagacagcag ctttggcaat ggagccattc caccctatgg tgaatctgga ttgttctcgg
     6001 gatttccggc cttttctttg tgcactctac gctcctattt gtatggaata tggacgtgtc
     6061 acacttccct gtcgtaggct gtgtcagcgg gcttacagtg agtgttcgaa gctcatggag
     6121 atgtttggtg ttccttggcc tgaagatatg gaatgcagta ggttcccaga ttgtgatgag
     6181 ccatatcctc gacttgtgga tctgaattta gctggagaac caactgaagg agccccagtg
     6241 gcagtgcaga gagactatgg tttttggtgt ccccgagagt taaaaattga tcctgatctg
     6301 ggttattctt ttctgcatgt gcgtgattgt tcacctcctt gtggctacga tgctggcttg
     6361 tacagccgct cagctaagga gttcacggat atttggatgg ctgtgtgggc cagcctctgc
     6421 ttcatctcca ccaccttcac cgtgctgacc ttcctgattg attcatccag gttttcttac
     6481 cctgagcgcc ccatcatatt tctcagtatg tgctataata tttatagcat tgcttatatt
     6541 gttcggctga ctgtaggccg ggaaaggata tcctgtgatt ttgaagaggc ggcagagccc
     6601 gttctcatcc aagaaggact taagaacaca ggatgtgcaa taattttctt gctgatgtac
     6661 ttttttggaa tggccagctc catttggtgg gttattctga cactcacttg gtttttggca
     6721 gccggactca agtggggtca tgaagccatt gaaatgcaca gttcttattt ccacatcgca
     6781 gcctgggcta ttcccgcagt gaaaaccatt gtcatcttga ttatgagact agtggatgcc
     6841 gatgaactga ctggcttgtg ctatgttggg aaccaaaacc tagatgccct cactggcttt
     6901 gtggtggctc ctctctttac gtatttggtg attggaacgc tgttcattgc ggcgggt
//
