LOCUS       sequence-256368-        8533 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Piggybac vector for constitutive expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8533)
  AUTHORS   Fischer AD, Veronese-Paniagua DA, Swaminathan S, Kashima H, Rubin 
            DC, Madison BB
  TITLE     The oncogenic function of PLAGL2 is mediated via ASCL2 and IGF2 and 
            a Wnt-independent mechanism in colorectal cancer.
  JOURNAL   Am J Physiol Gastrointest Liver Physiol. 2023 Jun 13. doi: 
            10.1152/ajpgi.00058.2022.
  PUBMED    37310750
REFERENCE   2  (bases 1 to 8533)
  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: "Am J 
            Physiol Gastrointest Liver Physiol. 2023 Jun 13. doi: 
            10.1152/ajpgi.00058.2022."
COMMENT     Sequence Label: BII-ChPtW-PLAGL2
FEATURES             Location/Qualifiers
     source          1..8533
                     /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)"
     primer_bind     complement(308..324)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(308..324)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     CDS             1875..1928
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             1935..1961
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             1935..1961
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     misc_feature    3510..4098
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3563..3583)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3981..3992)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4178..4312
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4202..4221)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4256..4275
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   5090..5152
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(5282..5299)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5282..5298)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5291..5313)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5440..5895
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5527..5546)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5737..5758
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5921..6025
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6026..6094,6095..6886)
                     /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(6244..6263)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7057..7645
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7546..7565
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7799..7816
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7933..7954
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7969..7974,7975..7992,7993..7999)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8007..8023
                     /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     8012..8034
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     8031..8047
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     8031..8047
                     /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 ccactagttc tagagggaca gccccccccc aaagccccca gggatgtaat
      361 tacgtccctc ccccgctagg gggcagcagc gagccgcccg gggctccgct ccggtccggc
      421 gctccccccg catccccgag ccggcagcgt gcggggacag cccgggcacg gggaaggtgg
      481 cacgggatcg ctttcctctg aacgcttctc gctgctcttt gagcctgcag acacctgggg
      541 ggatacgggg aaaaggcctc caaggcctgc aggagtcaat gggaaaaacc cattggagcc
      601 aagtacactg actcaatagg gactttccat tgggttttgc ccagtacata aggtcaatag
      661 ggggtgagtc aacaggaaag tcccattgga gccaagtaca ttgagtcaat agggactttc
      721 caatgggttt tgcccagtac ataaggtcaa tgggaggtaa gccaatgggt ttttcccatt
      781 actgacatgt atactgagtc attagggact ttccaatggg ttttgcccag tacataaggt
      841 caataggggt gaatcaacag gaaagtccca ttggagccaa gtacactgag tcaataggga
      901 ctttccattg ggttttgccc agtacaaaag gtcaataggg ggtgagtcaa tgggtttttc
      961 ccattattgg cacatacata aggtcaatag gggtgactag tggagaagag catgcttgag
     1021 ggctgagtgc ccctcagtgg gcagagagca catggcccac agtccctgag aagttggggg
     1081 gaggggtggg caattgaact ggtgcctaga gaaggtgggg cttgggtaaa ctgggaaagt
     1141 gatgtggtgt actggctcca cctttttccc cagggtgggg gagaaccata tataagtgca
     1201 gtagtctctg tgaacattcg gtttagtgaa cctgcaggtc ggccgccacg accggtgcca
     1261 ccatgactga atacaaacca actgttcgcc tggcaactcg tgatgatgtt ccacgtgcag
     1321 ttcgcaccct ggctgctgca tttgctgact accctgcaac ccgtcacact gtggacccag
     1381 accgccacat tgaacgtgtg actgaactgc aggagctgtt cctgacccgt gtgggcctgg
     1441 acattggcaa agtgtgggtg gcagatgatg gtgctgctgt ggcagtgtgg accacccctg
     1501 aatctgttga agctggtgca gtgtttgctg agattggccc acgcatggca gaactgtctg
     1561 gcagccgcct ggcagcacaa cagcagatgg aaggtctgct ggcaccacac cgcccaaaag
     1621 aacctgcttg gttcctggca actgtgggtg tgagccctga ccaccagggt aagggcctgg
     1681 gctctgcagt ggtgctgcct ggtgtggaag cagctgaacg tgcaggtgtg cctgctttcc
     1741 tggagacctc agctccacgc aacctgcctt tctatgaacg cctgggcttc actgtgactg
     1801 ctgatgtgga agtgccagaa ggcccacgca cttggtgcat gactcgcaaa ccaggtgcta
     1861 gccttggctc cggcgagggc aggggaagtc ttctaacatg cggggacgtg gaggaaaatc
     1921 ccggcccact cggctacccc tacgacgtgc ccgactacgc cagcggcatg accacatttt
     1981 tcaccagcgt ccccccctgg attcaagatg caaagcagga ggaggaagtg ggctggaaac
     2041 tagttcccag gcctcggggc cgggaggcgg agagtcaagt gaagtgccaa tgtgaaattt
     2101 cgggaacacc tttctcaaat ggggagaagc tgaggcctca cagcctcccg caaccagagc
     2161 agagaccata tagctgccct cagctgcact gtggcaaggc ttttgcttcc aaatacaagc
     2221 tgtataggca catggccacc cactcagccc agaaacccca ccagtgtatg tactgtgata
     2281 agatgtttca ccgcaaggac catctgcgga accatctgca gacccatgat cctaacaaag
     2341 aggccctcca ctgctctgag tgcggtaaga attacaatac gaagctgggc taccggcgcc
     2401 acctggccat gcatgctgcc agcagcggtg acctcagctg caaggtgtgc ctgcagacct
     2461 ttgagagtac ccaggccctg ctagagcacc tgaaggccca ctcacgccgg gtagcaggcg
     2521 gtgccaagga gaagaagcac ccctgtgacc actgcgaccg gcggttctat actcgtaagg
     2581 atgtacggcg gcacctagtg gtgcacacag gccgtaagga cttcctgtgc cagtactgtg
     2641 cccagcggtt tggccgtaag gaccacctga cgcgtcatgt caagaagagc cactcgcagg
     2701 agctgctcaa gatcaagaca gagcccgtgg acatgttagg cctactcagc tgcagctcca
     2761 cagtcagtgt gaaggaagag ctgagccctg tgctgtgcat ggcctctcgg gacgtaatgg
     2821 ggaccaaggc cttccctggc atgttgccca tgggcatgta tggtgcccac atccctacca
     2881 tgcccagcac gggcgtgcca cactccctgg tgcacaacac gctgcccatg ggtatgagct
     2941 accctctgga atcctcacct atctcttccc cagctcagct ccctccaaaa taccagcttg
     3001 gatctacctc atacttgccc gacaaattgc ccaaagtgga ggtggatagt tttctggcgg
     3061 agcttcctgg aagcctgtct ctctcatccg ctgaacccca gcccgcctca cctcagccgg
     3121 cggcagctgc ggccctccta gatgaagcac tgcttgccaa gagccccgcc aacctctctg
     3181 aggccctctg cgctgctaat gtggacttct cccacctact gggctttctt ccactcaacc
     3241 tgcccccgtg taacccacct ggggccacag gaggcctggt catgggctac tcccaggctg
     3301 aggcacagcc cctgcttacc actttgcaag ctcagcctca agattcccca ggagctgggg
     3361 gaccactgaa ctttgggcct ctgcactcct tgcctcctgt cttcacgtct ggcctgagta
     3421 gcaccaccct gcctcgtttc catcaagcat tccagtagcc cccacagagg ccctcagctc
     3481 agtttttggc tagagttcgc gtctggaaca atcaacctct ggattacaaa atttgtgaaa
     3541 gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa
     3601 tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat
     3661 cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt
     3721 gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc
     3781 tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc
     3841 ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg
     3901 ggaagctgac gtcctttcca tggctgctcg cctgtgttgc cacctggatt ctgcgcggga
     3961 cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc
     4021 tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc
     4081 tttgggccgc ctccccgcct ggaattaatt ctggctaagc acttcgtggc cgtcgatcgt
     4141 ttaaagggag gtagtgagtc gagatatcga tactagtgat ccagacatga taagatacat
     4201 tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta tttgtgaaat
     4261 ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag ttaacaacaa
     4321 caattgcatt cattttatgt ttcaggttca gggggaggtg tgggaggttt tttaaagcaa
     4381 gtaaaacctc tacaaatgtg gtatggctga ttatgatcct ctagagggac agcccccccc
     4441 caaagccccc agggatgtaa ttacgtccct cccccgctag ggggcagcag cgagccgccc
     4501 ggggctccgc tccggtccgg cgctcccccc gcatccccga gccggcagcg tgcggggaca
     4561 gcccgggcac ggggaaggtg gcacgggatc gctttcctct gaacgcttct cgctgctctt
     4621 tgagcctgca gacacctggg gggatacggg gaaaagccat accaatggag ctcacgggga
     4681 cagccccccc ccaaagcccc cagggatgta attacgtccc tcccccgcta gggggcagca
     4741 gcgagccgcc cggggctccg ctccggtccg gcgctccccc cgcatccccg agccggcagc
     4801 gtgcggggac agcccgggca cggggaaggt ggcacgggat cgctttcctc tgaacgcttc
     4861 tcgctgctct ttgagcctgc agacacctgg ggggatacgg ggaaaaagaa ttcgataaaa
     4921 gttttgttac tttatagaag aaattttgag tttttgtttt ttttaataaa taaataaaca
     4981 taaataaatt gtttgttgaa tttattatta gtatgtaagt gtaaatataa taaaacttaa
     5041 tatctattca aattaataaa taaacctcga tatacagacc gataaaacac atgcgtcaat
     5101 tttacacatg attatcttta acgtacgtca caatatgatt atctttctag ggttaatcta
     5161 gctgcgtgtt ctgcagcgtg tcgagcatct tcatctgctc catcacgctg taaaacacat
     5221 ttgcaccgcg agtctgcccg tcctccacgg gttcaaaaac gtgaatgaac gaggcgcgct
     5281 cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc
     5341 gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc
     5401 gcccttccca acagttgcgc agcctgaatg gcgaatggga cgcgccctgt agcggcgcat
     5461 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag
     5521 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc
     5581 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc
     5641 ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt
     5701 ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa
     5761 caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg
     5821 cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat
     5881 taacgcttac aatttaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     5941 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     6001 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     6061 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     6121 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     6181 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     6241 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     6301 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     6361 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     6421 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     6481 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     6541 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     6601 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     6661 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     6721 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     6781 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     6841 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     6901 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     6961 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     7021 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     7081 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     7141 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     7201 tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     7261 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     7321 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     7381 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     7441 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     7501 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     7561 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     7621 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     7681 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     7741 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     7801 cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg
     7861 ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga
     7921 gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat
     7981 gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag
     8041 ctatgaccat gattacgcca agcgcgcccg ccgggtaact cacggggtat ccatgtccat
     8101 ttctgcggca tccagccagg atacccgtcc tcgctgacgt aatatcccag cgccgcaccg
     8161 ctgtcattaa tctgcacacc ggcacggcag ttccggctgt cgccggtatt gttcgggttg
     8221 ctgatgcgct tcgggctgac catccggaac tgtgtccgga aaagccgcga cgaactggta
     8281 tcccaggtgg cctgaacgaa cagttcaccg ttaaaggcgt gcatggccac accttcccga
     8341 atcatcatgg taaacgtgcg ttttcgctca acgtcaatgc agcagcagtc atcctcggca
     8401 aactctttcc atgccgcttc aacctcgcgg gaaaaggcac gggcttcttc ctccccgatg
     8461 cccagatagc gccagcttgg gcgatgactg agccggaaaa aagacccgac gatatgatcc
     8521 tgatgcagct aga
//
