LOCUS       sequence-215786-        7615 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of destabilized GFP under the control of the M38-TOP 
            canonical Wnt signaling responsive promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7615)
  AUTHORS   Boitard M, Bocchi R, Egervari K, Petrenko V, Viale B, Gremaud S, 
            Zgraggen E, Salmon P, Kiss JZ
  TITLE     Wnt signaling regulates multipolar-to-bipolar transition of 
            migrating neurons in the cerebral cortex.
  JOURNAL   Cell Rep. 2015 Mar 3;10(8):1349-61. doi: 
            10.1016/j.celrep.2015.01.061. Epub 2015 Feb 26.
  PUBMED    25732825
REFERENCE   2  (bases 1 to 7615)
  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"; doi: 
            "10.1016/j.celrep.2015.01.061"; journalName: "Cell Rep"; date: 
            "2015-03-3- 3"; volume: "10"; issue: "8"; pages: "1349-61"
COMMENT     Sequence Label: pCLX-M38s/TOP-dGFP
FEATURES             Location/Qualifiers
     source          1..7615
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     LTR             385..642
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    689..814
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1306..1423
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     misc_feature    1783..2016
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2201..2245
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     protein_bind    2522..2542
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     protein_bind    2719..2743
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             join(2749..2751,2752..2754,2755..3465,3466..3471,
                     3472..3594)
                     /codon_start=1
                     /product="EGFP destabilized by fusion to residues 422-461 
                     of mouse ornithine decarboxylase, giving an in vivo 
                     half-life of ~2 hours"
                     /label=d2EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYKKLSHGFPPEVEEQDDGTLPMSCAQESGMDRHPAACASARINV
                     "
     primer_bind     complement(2794..2815)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3055..3074)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3402..3423
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(3595..3619)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    3637..4225
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3690..3710)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4108..4119)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     protein_bind    complement(4368..4401)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     LTR             4421..4601
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(4639..4658)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4704..4759
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4758..4777)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(5118..5134)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5118..5134)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5131..5153)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5142..5158
                     /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(5166..5172,5173..5190,5191..5196))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5211..5232
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5291..5486
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5337..5472
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5492..5626
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5529..5548)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5583..5602
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5694..5711)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5865..6453)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5945..5964)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6624..7415,7416..7484))
                     /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     7247..7266
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7485..7589)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggact atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atgggaggcg tggcctgggc gggactgggg agtggcgagc
      421 cctcagatcc tgcatataag cagctgcttt ttgcctgtac tgggtctctc tggttagacc
      481 agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa
      541 gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga
      601 gatccctcag acccttttag tcagtgtgga aaatctctag cagtggcgcc cgaacaggga
      661 cttgaaagcg aaagggaaac cagaggagct ctctcgacgc aggactcggc ttgctgaagc
      721 gcgcacggca agaggcgagg ggcggcgact ggtgagtacg ccaaaaattt tgactagcgg
      781 aggctagaag gagagagatg ggtgcgagag cgtcagtatt aagcggggga gaattagatc
      841 gcgatgggaa aaaattcggt taaggccagg gggaaagaaa aaatataaat taaaacatat
      901 agtatgggca agcagggagc tagaacgatt cgcagttaat cctggcctgt tagaaacatc
      961 agaaggctgt agacaaatac tgggacagct acaaccatcc cttcagacag gatcagaaga
     1021 acttagatca ttatataata cagtagcaac cctctattgt gtgcatcaaa ggatagagat
     1081 aaaagacacc aaggaagctt tagacaagat agaggaagag caaaacaaaa gtaagaaaaa
     1141 agcacagcaa gcagcaggat cttcagacct ggaaattccc tacaatcccc aaagtcaagg
     1201 agtagtagaa tctatgaata aagaattaaa gaaaattata ggacaggtaa gagatcaggc
     1261 tgaacatctt aagacagcag tacaaatggc agtattcatc cacaatttta aaagaaaagg
     1321 ggggattggg gggtacagtg caggggaaag aatagtagac ataatagcaa cagacataca
     1381 aactaaagaa ttacaaaaac aaattacaaa aattcaaaat tttcgggttt attacaggga
     1441 cagcagaaat ccactttgga aaggaccagc aaagctcctc tggaaaggtg aaggggcagt
     1501 agtaatacaa gataatagtg acataaaagt agtgccaaga agaaaagcaa agatcattag
     1561 ggattatgga aaacagatgg caggtgatga ttgtgtggca agtagacagg atgaggatta
     1621 gaacatggaa aagtttagta aaacaccata aggaggagat atgagggaca attggagaag
     1681 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc
     1741 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg
     1801 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc
     1861 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc
     1921 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct
     1981 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa
     2041 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca
     2101 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt
     2161 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt
     2221 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta
     2281 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt
     2341 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct
     2401 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga
     2461 cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcgctagt gattatcgga
     2521 tcaactttgt atagaaaagt tgctgcagcc caagctatca aagggggtaa gatcaaaggg
     2581 ggtaagatca aagggggtaa gatcaaagga ggagcttcag tcgaccccag tgacgtagga
     2641 agtccatcca ttcacagcgc ttctataaag gcgccagctg aggcgcctac tactccaacc
     2701 gcgactgcag cgagcaactc aagtttgtac aaaaaagcag gcttcaccat ggtgagcaag
     2761 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
     2821 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
     2881 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
     2941 ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc
     3001 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
     3061 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
     3121 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
     3181 aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg
     3241 aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag
     3301 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc
     3361 cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
     3421 gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagaagct tagccatggc
     3481 ttcccgccgg aggtggagga gcaggatgat ggcacgctgc ccatgtcttg tgcccaggag
     3541 agcgggatgg accgtcaccc tgcagcctgt gcttctgcta ggatcaatgt gtagacccag
     3601 ctttcttgta caaagtggtg ataatcgaat tccgataatc aacctctgga ttacaaaatt
     3661 tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct
     3721 gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg
     3781 tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc
     3841 gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt
     3901 cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc
     3961 gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg
     4021 ttgtcgggga agctgacgtc ctttccatgg ctgctcgcct gtgttgccac ctggattctg
     4081 cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc
     4141 ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg
     4201 atctcccttt gggccgcctc cccgcatcgg gaattcccgc ggttcgcttt aagaccaatg
     4261 acttacaagg cagctgtaga tcttagccac tttttaaaag aaaagggggg actggaaggg
     4321 ctaattcact cccaacgaag acaagatggg atcaattcac catgggaata acttcgtata
     4381 gcatacatta tacgaagtta tgctgctttt tgcttgtact gggtctctct ggttagacca
     4441 gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag
     4501 cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag
     4561 atccctcaga cccttttagt cagtgtggaa aatctctagc agcatctagt tgaatcgcca
     4621 gtgtgtcgag ctagcagatc tttttccctc tgccaaaaat tatggggaca tcatgaagcc
     4681 ccttgagcat ctgacttctg gctaataaag gaaatttatt ttcattgcaa tagtgtgttg
     4741 gaattttttg tgtctctcac tcggaaggac atatgggagg gcaaatcatt taaaacatca
     4801 gaatgagtat ttggtttaga gtttggcaac atatgccata tgctggctgc catgaacaaa
     4861 ggtggctata aagaggtcat cagtatatga aacagccccc tgctgtccat tccttattcc
     4921 atagaaaagc cttgacttga ggttagattt tttttatatt ttgttttgtg ttattttttt
     4981 ctttaacatc cctaaaattt tccttacatg ttttactagc cagatttttc ctcctctcct
     5041 gactactccc agtcatagct gtccctcttc tcttatgaag atccctcgac ctgcagccca
     5101 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     5161 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     5221 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     5281 cagcggatcc gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     5341 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     5401 tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct
     5461 tttttggagg cctaggcttt tgcaaaaagc taacttgttt attgcagctt ataatggtta
     5521 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     5581 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggatccgct gcattaatga
     5641 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     5701 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     5761 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     5821 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     5881 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     5941 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     6001 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcaa
     6061 tgctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     6121 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     6181 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     6241 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     6301 agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     6361 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     6421 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     6481 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     6541 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     6601 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     6661 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     6721 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     6781 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     6841 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     6901 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     6961 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     7021 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     7081 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     7141 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     7201 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     7261 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     7321 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     7381 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     7441 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     7501 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     7561 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctg
//
