LOCUS       sequence-360422-        6976 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses FLAG tagged FZD4 swapped with FZD6 linker domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6976)
  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 6976)
  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_6linker
FEATURES             Location/Qualifiers
     source          1..6976
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             388..405
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(428..445)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    434..658
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      704..1132
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(791..810)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1001..1022
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1141..1161)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1146..1475
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1326..1461
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1388..1407
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             1542..2336
                     /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(1596..1615)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2206..2225
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    2512..2633
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2549..2568)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2603..2622
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2682..2698)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2682..2698)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2695..2717)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2706..2722
                     /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(2730..2736,2737..2754,2755..2760))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2775..2796
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2913..2930)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3084..3672)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3164..3183)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3843..4634,4635..4703))
                     /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     4466..4485
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4704..4808)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4883..4902)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        5074..5453
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5454..5657
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5608..5628
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5702..5721
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5702..5720
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5753..5797
                     /codon_start=1
                     /product="leader sequence from mouse preprotrypsin"
                     /label=preprotrypsin leader
                     /translation="MSALLILALVGAAVA"
     CDS             5798..5821
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
ORIGIN
        1 gccttattca aaattcggtc caatcttcaa aaagacggga caaagacaga caagttggaa
       61 aggctaatgg tcaagatcgg ggtcttctca gtactgtaca cggttcctgc aacctgtgtg
      121 attgcctgtt atttctatga aatctcaaac tgggcactct ttcgatattc tgcagatgac
      181 tcaaacatgg cagttgaaat gttgaaaatt tttatgtctt tgctcgtggg catcacttca
      241 ggcatgtgga tttggtctgc caaaactctt cacacgtggc aaaagtgttc taaccgattg
      301 gtgaattctg ggaaggtaaa gagagagaag agggggaatg gttgggtgaa gccaggaaaa
      361 ggcaacgaga ctgtggtata gctcgagcat catcaccatc accattgagt ttaaacccgc
      421 tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
      481 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt
      541 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc
      601 aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct
      661 tctgaggcgg aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc
      721 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
      781 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
      841 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
      901 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
      961 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     1021 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     1081 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt
     1141 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     1201 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     1261 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     1321 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     1381 ttttttttat ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt
     1441 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca
     1501 ttttcggatc tgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat
     1561 tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac
     1621 agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc
     1681 tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaggacgag gcagcgcggc
     1741 tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag
     1801 cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc
     1861 ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg
     1921 atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc
     1981 ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc
     2041 cagccgaact gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga
     2101 cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca
     2161 tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg
     2221 atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg
     2281 ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg
     2341 gactctgggg ttcgcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc
     2401 gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc
     2461 tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt
     2521 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
     2581 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc
     2641 tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg
     2701 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     2761 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     2821 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     2881 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     2941 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     3001 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     3061 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     3121 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     3181 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     3241 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     3301 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     3361 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     3421 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     3481 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     3541 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     3601 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     3661 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     3721 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     3781 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     3841 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     3901 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     3961 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     4021 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     4081 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     4141 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     4201 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     4261 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     4321 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     4381 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     4441 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     4501 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     4561 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     4621 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     4681 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     4741 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     4801 gttccgcgca catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctcccg
     4861 atcccctatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtatc
     4921 tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa
     4981 caaggcaagg cttgaccgac aattgcatga agaatctgct tagggttagg cgttttgcgc
     5041 tgcttcgcga tgtacgggcc agatatacgc gttgacattg attattgact agttattaat
     5101 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac
     5161 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa
     5221 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
     5281 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc
     5341 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat
     5401 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc
     5461 ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc
     5521 tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa
     5581 aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg
     5641 tctatataag cagagctctc tggctaacta gagaacccac tgcttactgg cttatcgaaa
     5701 ttaatacgac tcactatagg gagacccaag ctggctagtt aagcttggta ccatgtctgc
     5761 acttctgatc ctagctcttg ttggagctgc agttgctgac tacaaagacg atgacgacaa
     5821 gcttgcggcc gcagaggagc ggcgctgcga ccccatccgc atcgccatgt gccagaacct
     5881 cggctacaac gtgaccaaga tgcccaactt agtgggacac gagctgcaga cagacgccga
     5941 gctgcagctg acaactttca cgccgctcat ccagtacggc tgctccagcc agctgcagtt
     6001 cttcctttgt tcggtttatg tgccaatgtg cacagagaag atcaacatcc ccatcggccc
     6061 gtgcggtggc atgtgccttt cagtcaagag acgctgtgaa ccagtcctga gagaatttgg
     6121 gtttgcctgg cccgacaccc tgaactgcag caagttcccg ccccagaacg accacaacca
     6181 catgtgcatg gaaactgttc ctgtaacttt tgatccacac acagaatttc ttggtcctca
     6241 gaagaaaaca gaacaagtcc aaagagacat tggattttgg tgtccaaggc atcttaagac
     6301 ttctggggga caaggatata agtttctggg aattgaccag tgtgcgcctc catgtggcta
     6361 cgatgctggc ttgtacagcc gctcagctaa ggagttcacg gatatttgga tggctgtgtg
     6421 ggccagcctc tgcttcatct ccaccacctt caccgtgctg accttcctga ttgattcatc
     6481 caggttttct taccctgagc gccccatcat atttctcagt atgtgctata atatttatag
     6541 cattgcttat attgttcggc tgactgtagg ccgggaaagg atatcctgtg attttgaaga
     6601 ggcggcagag cccgttctca tccaagaagg acttaagaac acaggatgtg caataatttt
     6661 cttgctgatg tacttttttg gaatggccag ctccatttgg tgggttattc tgacactcac
     6721 ttggtttttg gcagccggac tcaagtgggg tcatgaagcc attgaaatgc acagttctta
     6781 tttccacatc gcagcctggg ctattcccgc agtgaaaacc attgtcatct tgattatgag
     6841 actagtggat gccgatgaac tgactggctt gtgctatgtt gggaaccaaa acctagatgc
     6901 cctcactggc tttgtggtgg ctcctctctt tacgtatttg gtgattggaa cgctgttcat
     6961 tgcggcgggt ttggtg
//
