LOCUS       sequence-256384-        7264 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Piggybac vector for canonical Wnt signaling reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7264)
  AUTHORS   Strubberg AM, Veronese Paniagua DA, Zhao T, Dublin L, Pritchard T, 
            Bayguinov PO, Fitzpatrick JAJ, Madison BB
  TITLE     The Zinc Finger Transcription Factor PLAGL2 Enhances Stem Cell Fate 
            and Activates Expression of ASCL2 in Intestinal Epithelial Cells.
  JOURNAL   Stem Cell Reports. 2018 Aug 14;11(2):410-424. doi: 
            10.1016/j.stemcr.2018.06.009. Epub 2018 Jul 12.
  PUBMED    30017821
REFERENCE   2  (bases 1 to 7264)
  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.stemcr.2018.06.009"; journalName: "Stem Cell Reports"; 
            date: "2018-08-14- 14"; volume: "11"; issue: "2"; pages: "410-424"
COMMENT     Sequence Label: BII-TOP-tdT
FEATURES             Location/Qualifiers
     source          1..7264
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   5..39
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     misc_feature    join(580..645,646..683)
                     /label=Ad SA
                     /note="adenovirus major late transcript splice acceptor 
                     (Friedrich and Soriano, 1991)"
     primer_bind     complement(703..722)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    747..795
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     polyA_signal    983..1031
                     /note="synthetic polyadenylation signal"
     misc_feature    1045..1136
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     1085..1104
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     promoter        1304..1335
                     /label=minP
                     /note="minimal TATA-box promoter with low basal activity"
     regulatory      1394..1403
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1400..2827
                     /codon_start=1
                     /product="tandem dimeric (pseudo-monomeric) derivative of 
                     DsRed (Shaner et al., 2004)"
                     /label=tdTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLGHGTGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPY
                     EGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVM
                     NFEDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGV
                     LKGEIHQALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYE
                     RSEGRHHLFLYGMDELYK"
     CDS             2831..2851
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    2914..3138
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3821..3883
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(4013..4030)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4013..4029)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4022..4044)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4171..4626
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4258..4277)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4468..4489
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4652..4756
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4757..4825,4826..5617)
                     /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(4975..4994)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5788..6376
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6277..6296
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6530..6547
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6664..6685
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6700..6705,6706..6723,6724..6730)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6738..6754
                     /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     6743..6765
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6762..6778
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6762..6778
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ttaaccctag aaagatagtc tgcgtaaaat tgacgcatgc attcttgaaa tattgctctc
       61 tctttctaaa tagcgcgaat ccgtcgctgt gcatttagga catctcagtc gccgcttgga
      121 gctcccgtga ggcgtgcttg tcaatgcggt aagtgtcact gattttgaac tataacgacc
      181 gcgtgagtca aaatgacgca tgattatctt ttacgtgact tttaagattt aactcatacg
      241 ataattatat tgttatttca tgttctactt acgtgataac ttattatata tatattttct
      301 tgttatagat ctgtggaatg tgtgctagag ggacagcccc cccccaaagc ccccagggat
      361 gtaattacgt ccctcccccg ctagggggca gcagcgagcc gcccggggct ccgctccggt
      421 ccggcgctcc ccccgcatcc ccgagccggc agcgtgcggg gacagcccgg gcacggggaa
      481 ggtggcacgg gatcgctttc ctctgaacgc ttctcgctgc tctttgagcc tgcagacacc
      541 tggggggata cggggaaaag gcctggtagg ctgcagcggt agggcgcagt agtccagggt
      601 ttccttgatg atgtcatact tatcctgtcc cttttttttc cacagctcgc ggttgaggac
      661 aaactcttcg cggtctttcc agtatgcatg ggggaggcta actgaaacac ggaaggagac
      721 aataccggaa ggaacccgcg ctatgacggc aataaaaaga cagaataaaa cgcacgggtg
      781 ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg ctggcactct gtcgataccc
      841 caccgagacc ccattggggc caatacgccc gcgtttcttc cttttcccca ccccaccccc
      901 caagttcggg tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc cctgccatag
      961 catattcggc cgccacggcc gcaataaaat atctttattt tcattacatc tgtgtgttgg
     1021 ttttttgtgt gaatcgatag tactaacata cgctctccat caaaacaaaa cgaaacaaaa
     1081 caaactagca aaataggctg tccccagtgc aagtgcaggt gccagaacat ttctctatcg
     1141 ataggtaccg agctcttacg cgagatcaaa gggggtaaga tcaaaggggg taagatcaaa
     1201 ggggcgcgag atcaaagggg gtaagatcaa agggggtaag atcaaagggg gtaagatcaa
     1261 aggggcgcgc ccgcgtgcta gcccgggctc gagatctaga ctctagaggg tatataatgg
     1321 aagctcgaat tccagcttgg cattccggta ctgttggtaa aaagcttggc attccggtac
     1381 tgttgccaac tcggccacca tggtgagcaa gggcgaggag gtcatcaaag agttcatgcg
     1441 cttcaaggtg cgcatggagg gctccatgaa cggccacgag ttcgagatcg agggcgaggg
     1501 cgagggccgc ccctacgagg gcacccagac cgccaagctg aaggtgacca agggcggccc
     1561 cctgcccttc gcctgggaca tcctgtcccc ccagttcatg tacggctcca aggcgtacgt
     1621 gaagcacccc gccgacatcc ccgattacaa gaagctgtcc ttccccgagg gcttcaagtg
     1681 ggagcgcgtg atgaacttcg aggacggcgg tctggtgacc gtgacccagg actcctccct
     1741 gcaggacggc acgctgatct acaaggtgaa gatgcgcggc accaacttcc cccccgacgg
     1801 ccccgtaatg cagaagaaga ccatgggctg ggaggcctcc accgagcgcc tgtacccccg
     1861 cgacggcgtg ctgaagggcg agatccacca ggccctgaag ctgaaggacg gcggccacta
     1921 cctggtggag ttcaagacca tctacatggc caagaagccc gtgcaactgc ccggctacta
     1981 ctacgtggac accaagctgg acatcacctc ccacaacgag gactacacca tcgtggaaca
     2041 gtacgagcgc tccgagggcc gccaccacct gttcctgggg catggcaccg gcagcaccgg
     2101 cagcggcagc tccggcaccg cctcctccga ggacaacaac atggccgtca tcaaagagtt
     2161 catgcgcttc aaggtgcgca tggagggctc catgaacggc cacgagttcg agatcgaggg
     2221 cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg
     2281 cggccccctg cccttcgcct gggacatcct gtccccccag ttcatgtacg gctccaaggc
     2341 gtacgtgaag caccccgccg acatccccga ttacaagaag ctgtccttcc ccgagggctt
     2401 caagtgggag cgcgtgatga acttcgagga cggcggtctg gtgaccgtga cccaggactc
     2461 ctccctgcag gacggcacgc tgatctacaa ggtgaagatg cgcggcacca acttcccccc
     2521 cgacggcccc gtaatgcaga agaagaccat gggctgggag gcctccaccg agcgcctgta
     2581 cccccgcgac ggcgtgctga agggcgagat ccaccaggcc ctgaagctga aggacggcgg
     2641 ccactacctg gtggagttca agaccatcta catggccaag aagcccgtgc aactgcccgg
     2701 ctactactac gtggacacca agctggacat cacctcccac aacgaggact acaccatcgt
     2761 ggaacagtac gagcgctccg agggccgcca ccacctgttc ctgtacggca tggacgagct
     2821 gtacaagggc cccaagaaga agcgcaaggt gggcgcctaa gctaagcact tcgtggccgt
     2881 cgatcgttta aagggaggta gtgagtcgag cgactgtgcc ttctagttgc cagccatctg
     2941 ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt
     3001 cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg
     3061 gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg
     3121 atgcggtggg ctctatggcc tgttaggcct tctagaggga cagccccccc ccaaagcccc
     3181 cagggatgta attacgtccc tcccccgcta gggggcagca gcgagccgcc cggggctccg
     3241 ctccggtccg gcgctccccc cgcatccccg agccggcagc gtgcggggac agcccgggca
     3301 cggggaaggt ggcacgggat cgctttcctc tgaacgcttc tcgctgctct ttgagcctgc
     3361 agacacctgg ggggatacgg ggaaaagcca taccaatgga gctcacgggg acagcccccc
     3421 cccaaagccc ccagggatgt aattacgtcc ctcccccgct agggggcagc agcgagccgc
     3481 ccggggctcc gctccggtcc ggcgctcccc ccgcatcccc gagccggcag cgtgcgggga
     3541 cagcccgggc acggggaagg tggcacggga tcgctttcct ctgaacgctt ctcgctgctc
     3601 tttgagcctg cagacacctg gggggatacg gggaaaaaga attcgataaa agttttgtta
     3661 ctttatagaa gaaattttga gtttttgttt tttttaataa ataaataaac ataaataaat
     3721 tgtttgttga atttattatt agtatgtaag tgtaaatata ataaaactta atatctattc
     3781 aaattaataa ataaacctcg atatacagac cgataaaaca catgcgtcaa ttttacacat
     3841 gattatcttt aacgtacgtc acaatatgat tatctttcta gggttaatct agctgcgtgt
     3901 tctgcagcgt gtcgagcatc ttcatctgct ccatcacgct gtaaaacaca tttgcaccgc
     3961 gagtctgccc gtcctccacg ggttcaaaaa cgtgaatgaa cgaggcgcgc tcactggccg
     4021 tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag
     4081 cacatccccc tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc
     4141 aacagttgcg cagcctgaat ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg
     4201 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     4261 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     4321 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     4381 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     4441 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     4501 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     4561 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta
     4621 caatttaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     4681 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     4741 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     4801 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     4861 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     4921 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     4981 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     5041 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     5101 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     5161 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     5221 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     5281 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
     5341 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     5401 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     5461 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     5521 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     5581 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     5641 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     5701 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     5761 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     5821 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     5881 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct
     5941 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     6001 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     6061 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     6121 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     6181 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     6241 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     6301 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     6361 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     6421 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     6481 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     6541 gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat
     6601 tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc
     6661 aattaatgtg agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc
     6721 tcgtatgttg tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacca
     6781 tgattacgcc aagcgcgccc gccgggtaac tcacggggta tccatgtcca tttctgcggc
     6841 atccagccag gatacccgtc ctcgctgacg taatatccca gcgccgcacc gctgtcatta
     6901 atctgcacac cggcacggca gttccggctg tcgccggtat tgttcgggtt gctgatgcgc
     6961 ttcgggctga ccatccggaa ctgtgtccgg aaaagccgcg acgaactggt atcccaggtg
     7021 gcctgaacga acagttcacc gttaaaggcg tgcatggcca caccttcccg aatcatcatg
     7081 gtaaacgtgc gttttcgctc aacgtcaatg cagcagcagt catcctcggc aaactctttc
     7141 catgccgctt caacctcgcg ggaaaaggca cgggcttctt cctccccgat gcccagatag
     7201 cgccagcttg ggcgatgact gagccggaaa aaagacccga cgatatgatc ctgatgcagc
     7261 taga
//
