LOCUS       sequence-218672-        7616 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 7616)
  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 7616)
  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..7616
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     145..166
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(338..362)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    380..968
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(433..453)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(851..862)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     protein_bind    complement(1111..1144)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     LTR             1164..1344
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(1382..1401)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    1447..1502
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(1501..1520)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(1863..1879)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1863..1879)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1876..1898)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1887..1903
                     /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(1911..1917,1918..1935,1936..1941))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1956..1977
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        2036..2231
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      2082..2217
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    2237..2371
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2274..2293)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2328..2347
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2439..2456)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2610..3198)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2690..2709)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3369..4160,4161..4229))
                     /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     3992..4011
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4230..4334)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        4365..4744
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     LTR             4746..5003
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5050..5175
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    5667..5784
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     misc_feature    6144..6377
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6562..6606
                     /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    6881..6901
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     protein_bind    7078..7102
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             join(7108..7110,7111..7113,7114..7616,1..208,209..214,
                     215..337)
                     /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(7153..7174)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(7414..7433)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
ORIGIN
        1 caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg
       61 cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa
      121 agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat
      181 cactctcggc atggacgagc tgtacaagaa gcttagccat ggcttcccgc cggaggtgga
      241 ggagcaggat gatggcacgc tgcccatgtc ttgtgcccag gagagcggga tggaccgtca
      301 ccctgcagcc tgtgcttctg ctaggatcaa tgtgtagacc cagctttctt gtacaaagtg
      361 gtgataatcg aattccgata atcaacctct ggattacaaa atttgtgaaa gattgactgg
      421 tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta
      481 tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct
      541 gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt
      601 tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac
      661 tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg
      721 ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaagctgac
      781 gtcctttcca tggctgctcg cctgtgttgc cacctggatt ctgcgcggga cgtccttctg
      841 ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct
      901 gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc
      961 ctccccgcat cgggaattcc cgcggttcgc tttaagacca atgacttaca aggcagctgt
     1021 agatcttagc cactttttaa aagaaaaggg gggactggaa gggctaattc actcccaacg
     1081 aagacaagat gggatcaatt caccatggga ataacttcgt atagcataca ttatacgaag
     1141 ttatgctgct ttttgcttgt actgggtctc tctggttaga ccagatctga gcctgggagc
     1201 tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct tgagtgcttc
     1261 aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc agaccctttt
     1321 agtcagtgtg gaaaatctct agcagcatct agttgaatcg ccagtgtgtc gagctagcag
     1381 atctttttcc ctctgccaaa aattatgggg acatcatgaa gccccttgag catctgactt
     1441 ctggctaata aaggaaattt attttcattg caatagtgtg ttggaatttt ttgtgtctct
     1501 cactcggaag gacatatggg agggcaaatc atttaaaaca tcagaatgag tatttggttt
     1561 agagtttggc aacatatgcc catatgctgg ctgccatgaa caaaggttgg ctataaagag
     1621 gtcatcagta tatgaaacag ccccctgctg tccattcctt attccataga aaagccttga
     1681 cttgaggtta gatttttttt atattttgtt ttgtgttatt tttttcttta acatccctaa
     1741 aattttcctt acatgtttta ctagccagat ttttcctcct ctcctgacta ctcccagtca
     1801 tagctgtccc tcttctctta tggagatccc tcgacctgca gcccaagctt ggcgtaatca
     1861 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     1921 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     1981 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagcg gatccgcatc
     2041 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     2101 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     2161 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     2221 gcttttgcaa aaagctaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     2281 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     2341 actcatcaat gtatcttatc atgtctggat ccgctgcatt aatgaatcgg ccaacgcgcg
     2401 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     2461 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     2521 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     2581 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     2641 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     2701 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     2761 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     2821 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     2881 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     2941 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     3001 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     3061 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     3121 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     3181 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     3241 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     3301 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     3361 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     3421 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     3481 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     3541 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     3601 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     3661 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     3721 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     3781 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     3841 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     3901 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     3961 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     4021 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     4081 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     4141 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     4201 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     4261 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     4321 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg ggtcgacatt gattattgac
     4381 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
     4441 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
     4501 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
     4561 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
     4621 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
     4681 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
     4741 catgggaggc gtggcctggg cgggactggg gagtggcgag ccctcagatc ctgcatataa
     4801 gcagctgctt tttgcctgta ctgggtctct ctggttagac cagatctgag cctgggagct
     4861 ctctggctaa ctagggaacc cactgcttaa gcctcaataa agcttgcctt gagtgcttca
     4921 agtagtgtgt gcccgtctgt tgtgtgactc tggtaactag agatccctca gaccctttta
     4981 gtcagtgtgg aaaatctcta gcagtggcgc ccgaacaggg acttgaaagc gaaagggaaa
     5041 ccagaggagc tctctcgacg caggactcgg cttgctgaag cgcgcacggc aagaggcgag
     5101 gggcggcgac tggtgagtac gccaaaaatt ttgactagcg gaggctagaa ggagagagat
     5161 gggtgcgaga gcgtcagtat taagcggggg agaattagat cgcgatggga aaaaattcgg
     5221 ttaaggccag ggggaaagaa aaaatataaa ttaaaacata tagtatgggc aagcagggag
     5281 ctagaacgat tcgcagttaa tactggcctg ttagaaacat cagaaggctg tagacaaata
     5341 ctgggacagc tacaaccatc ccttcagaca ggatcagaag aacttagatc attatataat
     5401 acagtagcaa ccctctattg tgtgcatcaa aggatagaga taaaagacac caaggaagct
     5461 ttagacaaga tagaggaaga gcaaaacaaa agtaagaaaa aagcacagca agcagcagga
     5521 tcttcagacc tggaaattcc ctacaatccc caaagtcaag gagtagtaga atctatgaat
     5581 aaagaattaa agaaaattat aggacaggta agagatcagg ctgaacatct taagacagca
     5641 gtacaaatgg cagtattcat ccacaatttt aaaagaaaag gggggattgg ggggtacagt
     5701 gcaggggaaa gaatagtaga cataatagca acagacatac aaactaaaga attacaaaaa
     5761 caaattacaa aaattcaaaa ttttcgggtt tattacaggg acagcagaaa tccactttgg
     5821 aaaggaccag caaagctcct ctggaaaggt gaaggggcag tagtaataca agataatagt
     5881 gacataaaag tagtgccaag aagaaaagca aagatcatta gggattatgg aaaacagatg
     5941 gcaggtgatg attgtgtggc aagtagacag gatgaggatt agaacatgga aaagtttagt
     6001 aaaacaccat aaggaggaga tatgagggac aattggagaa gtgaattata taaatataaa
     6061 gtagtaaaaa ttgaaccatt aggagtagca cccaccaagg caaagagaag agtggtgcag
     6121 agagaaaaaa gagcagtggg aataggagct ttgttccttg ggttcttggg agcagcagga
     6181 agcactatgg gcgcagcgtc aatgacgctg acggtacagg ccagacaatt attgtctggt
     6241 atagtgcagc agcagaacaa tttgctgagg gctattgagg cgcaacagca tctgttgcaa
     6301 ctcacagtct ggggcatcaa gcagctccag gcaagaatcc tggctgtgga aagataccta
     6361 aaggatcaac agctcctggg gatttggggt tgctctggaa aactcatttg caccactgct
     6421 gtgccttgga atgctagttg gagtaataaa tctctggaac agatttggaa tcacacgacc
     6481 tggatggagt gggacagaga aattaacaat tacacaagct taatacactc cttaattgaa
     6541 gaatcgcaaa accagcaaga aaagaatgaa caagaattat tggaattaga taaatgggca
     6601 agtttgtgga attggtttaa cataacaaat tggctgtggt atataaaatt attcataatg
     6661 atagtaggag gcttggtagg tttaagaata gtttttgctg tactttctat agtgaataga
     6721 gttaggcagg gatattcacc attatcgttt cagacccacc tcccaacccc gaggggaccc
     6781 gacaggcccg aaggaataga agaagaaggt ggagagagag acagagacag atccattcga
     6841 ttagtgaacg gatctcgacg gtatctagtg attatcggat caactttgta tagaaaagtt
     6901 gctgcagccc aagctatcaa agggggtaag atcaaagggg gtaagatcaa agggggtaag
     6961 atcaaaggag gagcttcagt cgaccccagt gacgtaggaa gtccatccat tcacagcgct
     7021 tctataaagg cgccagctga ggcgcctact actccaaccg cgactgcagc gagcaactca
     7081 agtttgtaca aaaaagcagg cttcaccatg gtgagcaagg gcgaggagct gttcaccggg
     7141 gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc
     7201 ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc
     7261 ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc
     7321 ttcagccgct accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa
     7381 ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc
     7441 gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc
     7501 aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc
     7561 tatatcatgg ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgcca
//
