LOCUS       sequence-267688-        7255 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 7255)
  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 7255)
  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..7255
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    22..70
                     /note="synthetic polyadenylation signal"
     misc_feature    84..175
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     124..143
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     promoter        343..374
                     /label=minP
                     /note="minimal TATA-box promoter with low basal activity"
     regulatory      433..442
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             439..1866
                     /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             1870..1890
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    1953..2177
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2851..2913
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(3043..3060)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3043..3059)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3052..3074)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3201..3656
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3288..3307)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3498..3519
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3682..3786
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3787..3855,3856..4647)
                     /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(4005..4024)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4818..5406
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5307..5326
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5560..5577
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5694..5715
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5730..5735,5736..5753,5754..5760)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5768..5784
                     /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     5773..5795
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5792..5808
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5792..5808
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   6299..6333
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     misc_feature    join(6874..6939,6940..6977)
                     /label=Ad SA
                     /note="adenovirus major late transcript splice acceptor 
                     (Friedrich and Soriano, 1991)"
     primer_bind     complement(6997..7016)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    7041..7089
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
ORIGIN
        1 atattcggcc gccacggccg caataaaata tctttatttt cattacatct gtgtgttggt
       61 tttttgtgtg aatcgatagt actaacatac gctctccatc aaaacaaaac gaaacaaaac
      121 aaactagcaa aataggctgt ccccagtgca agtgcaggtg ccagaacatt tctctatcga
      181 taggtaccga gctcttacgc gagatcaaag ggggtaagat caaagggggt aagatcaaag
      241 gggcgcgaga tcaaaggggg taagatcaaa gggggtaaga tcaaaggggg taagatcaaa
      301 ggggcgcgcc cgcgtgctag cccgggctcg agatctagac tctagagggt atataatgga
      361 agctcgaatt ccagcttggc attccggtac tgttggtaaa aagcttggca ttccggtact
      421 gttgccaact cggccaccat ggtgagcaag ggcgaggagg tcatcaaaga gttcatgcgc
      481 ttcaaggtgc gcatggaggg ctccatgaac ggccacgagt tcgagatcga gggcgagggc
      541 gagggccgcc cctacgaggg cacccagacc gccaagctga aggtgaccaa gggcggcccc
      601 ctgcccttcg cctgggacat cctgtccccc cagttcatgt acggctccaa ggcgtacgtg
      661 aagcaccccg ccgacatccc cgattacaag aagctgtcct tccccgaggg cttcaagtgg
      721 gagcgcgtga tgaacttcga ggacggcggt ctggtgaccg tgacccagga ctcctccctg
      781 caggacggca cgctgatcta caaggtgaag atgcgcggca ccaacttccc ccccgacggc
      841 cccgtaatgc agaagaagac catgggctgg gaggcctcca ccgagcgcct gtacccccgc
      901 gacggcgtgc tgaagggcga gatccaccag gccctgaagc tgaaggacgg cggccactac
      961 ctggtggagt tcaagaccat ctacatggcc aagaagcccg tgcaactgcc cggctactac
     1021 tacgtggaca ccaagctgga catcacctcc cacaacgagg actacaccat cgtggaacag
     1081 tacgagcgct ccgagggccg ccaccacctg ttcctggggc atggcaccgg cagcaccggc
     1141 agcggcagct ccggcaccgc ctcctccgag gacaacaaca tggccgtcat caaagagttc
     1201 atgcgcttca aggtgcgcat ggagggctcc atgaacggcc acgagttcga gatcgagggc
     1261 gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggc
     1321 ggccccctgc ccttcgcctg ggacatcctg tccccccagt tcatgtacgg ctccaaggcg
     1381 tacgtgaagc accccgccga catccccgat tacaagaagc tgtccttccc cgagggcttc
     1441 aagtgggagc gcgtgatgaa cttcgaggac ggcggtctgg tgaccgtgac ccaggactcc
     1501 tccctgcagg acggcacgct gatctacaag gtgaagatgc gcggcaccaa cttccccccc
     1561 gacggccccg taatgcagaa gaagaccatg ggctgggagg cctccaccga gcgcctgtac
     1621 ccccgcgacg gcgtgctgaa gggcgagatc caccaggccc tgaagctgaa ggacggcggc
     1681 cactacctgg tggagttcaa gaccatctac atggccaaga agcccgtgca actgcccggc
     1741 tactactacg tggacaccaa gctggacatc acctcccaca acgaggacta caccatcgtg
     1801 gaacagtacg agcgctccga gggccgccac cacctgttcc tgtacggcat ggacgagctg
     1861 tacaagggcc ccaagaagaa gcgcaaggtg ggcgcctaag ctaagcactt agtggccgtc
     1921 gatcgtttaa agggaggtag tgagtcgagc gactgtgcct tctagttgcc agccatctgt
     1981 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     2041 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     2101 tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga
     2161 tgcggtgggc tctatggctt ctagagggac agcccccccc caaagccccc agggatgtaa
     2221 ttacgtccct cccccgctag ggggcagcag cgagccgccc ggggctccgc tccggtccgg
     2281 cgctcccccc gcatccccga gccggcagcg tgcggggaca gcccgggcac ggggaaggtg
     2341 gcacgggatc gctttcctct gaacgcttct cgctgctctt tgagcctgca gacacctggg
     2401 gggatacggg gaaaaagcca taccaatgga gctcacgggg acagcccccc cccaaagccc
     2461 ccagggatgt aattacgtcc ctcccccgct agggggcagc agcgagccgc ccggggctcc
     2521 gctccggtcc ggcgctcccc ccgcatcccc gagccggcag cgtgcgggga cagcccgggc
     2581 acggggaagg tggcacggga tcgctttcct ctgaacgctt ctcgctgctc tttgagcctg
     2641 cagacacctg gggggatacg gggaaaaaga attcgataaa agttttgtta ctttatagaa
     2701 gaaattttga gtttttgttt tttttaataa ataaataaac ataaataaat tgtttgttga
     2761 atttattatt agtatgtaag tgtaaatata ataaaactta atatctattc aaattaataa
     2821 ataaacctcg atatacagac cgataaaaca catgcgtcaa ttttacacat gattatcttt
     2881 aacgtacgtc acaatatgat tatctttcta gggttaatct agctgcgtgt tctgcagcgt
     2941 gtcgagcatc ttcatctgct ccatcacgct gtaaaacaca tttgcaccgc gagtctgccc
     3001 gtcctccacg ggttcaaaaa cgtgaatgaa cgaggcgcgc tcactggccg tcgttttaca
     3061 acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc
     3121 tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg
     3181 cagcctgaat ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     3241 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     3301 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     3361 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     3421 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     3481 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
     3541 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
     3601 gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta caatttaggt
     3661 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     3721 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     3781 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     3841 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     3901 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     3961 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     4021 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     4081 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     4141 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     4201 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     4261 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     4321 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     4381 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     4441 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     4501 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     4561 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     4621 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     4681 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     4741 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     4801 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     4861 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     4921 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     4981 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     5041 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     5101 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     5161 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     5221 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     5281 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     5341 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     5401 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     5461 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     5521 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     5581 gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc
     5641 agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg
     5701 agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg
     5761 tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacca tgattacgcc
     5821 aagcgcgccc gccgggtaac tcacggggta tccatgtcca tttctgcggc atccagccag
     5881 gatacccgtc ctcgctgacg taatatccca gcgccgcacc gctgtcatta atctgcacac
     5941 cggcacggca gttccggctg tcgccggtat tgttcgggtt gctgatgcgc ttcgggctga
     6001 ccatccggaa ctgtgtccgg aaaagccgcg acgaactggt atcccaggtg gcctgaacga
     6061 acagttcacc gttaaaggcg tgcatggcca caccttcccg aatcatcatg gtaaacgtgc
     6121 gttttcgctc aacgtcaatg cagcagcagt catcctcggc aaactctttc catgccgctt
     6181 caacctcgcg ggaaaaggca cgggcttctt cctccccgat gcccagatag cgccagcttg
     6241 ggcgatgact gagccggaaa aaagacccga cgatatgatc ctgatgcagc tagattaacc
     6301 ctagaaagat agtctgcgta aaattgacgc atgcattctt gaaatattgc tctctctttc
     6361 taaatagcgc gaatccgtcg ctgtgcattt aggacatctc agtcgccgct tggagctccc
     6421 gtgaggcgtg cttgtcaatg cggtaagtgt cactgatttt gaactataac gaccgcgtga
     6481 gtcaaaatga cgcatgatta tcttttacgt gacttttaag atttaactca tacgataatt
     6541 atattgttat ttcatgttct acttacgtga taacttatta tatatatatt ttcttgttat
     6601 agatctgtgg aatgtgtgct agagggacag ccccccccca aagcccccag ggatgtaatt
     6661 acgtccctcc cccgctaggg ggcagcagcg agccgcccgg ggctccgctc cggtccggcg
     6721 ctccccccgc atccccgagc cggcagcgtg cggggacagc ccgggcacgg ggaaggtggc
     6781 acgggatcgc tttcctctga acgcttctcg ctgctctttg agcctgcaga cacctggggg
     6841 gatacgggga aaaggcctgg taggctgcag cggtagggcg cagtagtcca gggtttcctt
     6901 gatgatgtca tacttatcct gtcccttttt tttccacagc tcgcggttga ggacaaactc
     6961 ttcgcggtct ttccagtatg catgggggag gctaactgaa acacggaagg agacaatacc
     7021 ggaaggaacc cgcgctatga cggcaataaa aagacagaat aaaacgcacg ggtgttgggt
     7081 cgtttgttca taaacgcggg gttcggtccc agggctggca ctctgtcgat accccaccga
     7141 gaccccattg gggccaatac gcccgcgttt cttccttttc cccaccccac cccccaagtt
     7201 cgggtgaagg cccagggctc gcagccaacg tcggggcggc aggccctgcc atagc
//
