LOCUS       sequence-268590-        8375 bp    DNA     circular SYN 18-DEC-2025
DEFINITION  Piggybac vector for constitutive expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8375)
  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 8375)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 18, 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-ChiNW-IGF2
FEATURES             Location/Qualifiers
     source          1..8375
                     /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"
     misc_feature    join(1933..2507,2508..2510,2511..2519)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2100..2117)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2327..2346
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             2532..3326
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLAATGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2586..2605)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3196..3215
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     misc_feature    3351..3939
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3404..3424)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3822..3833)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4019..4153
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4043..4062)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4097..4116
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4932..4994
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(5124..5141)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5124..5140)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5133..5155)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5282..5737
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5369..5388)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5579..5600
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5763..5867
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5868..5936,5937..6728)
                     /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(6086..6105)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6899..7487
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7388..7407
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7641..7658
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7775..7796
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7811..7816,7817..7834,7835..7841)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7849..7865
                     /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     7854..7876
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7873..7889
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7873..7889
                     /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 accggtgcct
     1261 actctagagc tagcgtcgcc ggcttccaga caccaatggg aatcccaatg gggaagtcga
     1321 tgctggtgct tctcaccttc ttggccttcg cctcgtgctg cattgctgct taccgcccca
     1381 gtgagaccct gtgcggcggg gagctggtgg acaccctcca gttcgtctgt ggggaccgcg
     1441 gcttctactt cagcaggccc gcaagccgtg tgagccgtcg cagccgtggc atcgttgagg
     1501 agtgctgttt ccgcagctgt gacctggccc tcctggagac gtactgtgct acccccgcca
     1561 agtccgagag ggacgtgtcg acccctccga ccgtgcttcc ggacaacttc cccagatacc
     1621 ccgtgggcaa gttcttccaa tatgacacct ggaagcagtc cacccagcgc ctgcgcaggg
     1681 gcctgcctgc cctcctgcgt gcccgccggg gtcacgtgct cgccaaggag ctcgaggcgt
     1741 tcagggaggc caaacgtcac cgtcccctga ttgctctacc cacccaagac cccgcccacg
     1801 ggggcgcccc cccagagatg gccagcaatc ggaagtgagc aaaactgccg caagtctgca
     1861 gcccggcgcc accatcctgc agcctcctcc tgaccacgga cgtttcgaat ttaaatcgga
     1921 tccgcggccg cgcccctctc cctccccccc ccctaacgtt actggccgaa gccgcttgga
     1981 ataaggccgg tgtgcgtttg tctatatgtt attttccacc atattgccgt cttttggcaa
     2041 tgtgagggcc cggaaacctg gccctgtctt cttgacgagc attcctaggg gtctttcccc
     2101 tctcgccaaa ggaatgcaag gtctgttgaa tgtcgtgaag gaagcagttc ctctggaagc
     2161 ttcttgaaga caaacaacgt ctgtagcgac cctttgcagg cagcggaacc ccccacctgg
     2221 cgacaggtgc ctctgcggcc aaaagccacg tgtataagat acacctgcaa aggcggcaca
     2281 accccagtgc cacgttgtga gttggatagt tgtggaaaga gtcaaatggc tctcctcaag
     2341 cgtattcaac aaggggctga aggatgccca gaaggtaccc cattgtatgg gatctgatct
     2401 ggggcctcgg tgcacatgct ttacatgtgt ttagtcgagg ttaaaaaaac gtctaggccc
     2461 cccgaaccac ggggacgtgg ttttcctttg aaaaacacga tgataatatg gccacaacca
     2521 tggcgtccgg aatgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     2581 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     2641 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     2701 tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccgcgacg ggcgttcctt
     2761 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     2821 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     2881 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     2941 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     3001 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc
     3061 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     3121 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     3181 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
     3241 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     3301 atcgccttct tgacgagttc ttctgagtcg ctagagttcg cgtctggaac aatcaacctc
     3361 tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc
     3421 tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca
     3481 ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg
     3541 tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca
     3601 ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg
     3661 cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg
     3721 acaattccgt ggtgttgtcg gggaagctga cgtcctttcc atggctgctc gcctgtgttg
     3781 ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg
     3841 accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc
     3901 ctcagacgag tcggatctcc ctttgggccg cctccccgcc tggaattaat tctggctaag
     3961 cacttcgtgg ccgtcgatcg tttaaaggga ggtagtgagt cgagatatcg atactagtga
     4021 tccagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa
     4081 aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg
     4141 caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc agggggaggt
     4201 gtgggaggtt ttttaaagca agtaaaacct ctacaaatgt ggtatggctg attatgatcc
     4261 tctagaggga cagccccccc ccaaagcccc cagggatgta attacgtccc tcccccgcta
     4321 gggggcagca gcgagccgcc cggggctccg ctccggtccg gcgctccccc cgcatccccg
     4381 agccggcagc gtgcggggac agcccgggca cggggaaggt ggcacgggat cgctttcctc
     4441 tgaacgcttc tcgctgctct ttgagcctgc agacacctgg ggggatacgg ggaaaaagcc
     4501 ataccaatgg agctcacggg gacagccccc ccccaaagcc cccagggatg taattacgtc
     4561 cctcccccgc tagggggcag cagcgagccg cccggggctc cgctccggtc cggcgctccc
     4621 cccgcatccc cgagccggca gcgtgcgggg acagcccggg cacggggaag gtggcacggg
     4681 atcgctttcc tctgaacgct tctcgctgct ctttgagcct gcagacacct ggggggatac
     4741 ggggaaaaag aattcgataa aagttttgtt actttataga agaaattttg agtttttgtt
     4801 ttttttaata aataaataaa cataaataaa ttgtttgttg aatttattat tagtatgtaa
     4861 gtgtaaatat aataaaactt aatatctatt caaattaata aataaacctc gatatacaga
     4921 ccgataaaac acatgcgtca attttacaca tgattatctt taacgtacgt cacaatatga
     4981 ttatctttct agggttaatc tagctgcgtg ttctgcagcg tgtcgagcat cttcatctgc
     5041 tccatcacgc tgtaaaacac atttgcaccg cgagtctgcc cgtcctccac gggttcaaaa
     5101 acgtgaatga acgaggcgcg ctcactggcc gtcgttttac aacgtcgtga ctgggaaaac
     5161 cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat
     5221 agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg
     5281 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     5341 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     5401 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     5461 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     5521 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     5581 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     5641 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     5701 aacgcgaatt ttaacaaaat attaacgctt acaatttagg tggcactttt cggggaaatg
     5761 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     5821 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     5881 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     5941 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca
     6001 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc
     6061 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg
     6121 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac
     6181 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca
     6241 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg
     6301 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac
     6361 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg
     6421 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat
     6481 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg
     6541 ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg
     6601 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc
     6661 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc
     6721 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt
     6781 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     6841 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     6901 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     6961 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     7021 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     7081 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     7141 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     7201 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     7261 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     7321 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     7381 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     7441 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     7501 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     7561 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc
     7621 gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcccaatac
     7681 gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc
     7741 ccgactggaa agcgggcagt gagcgcaacg caattaatgt gagttagctc actcattagg
     7801 caccccaggc tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat
     7861 aacaatttca cacaggaaac agctatgacc atgattacgc caagcgcgcc cgccgggtaa
     7921 ctcacggggt atccatgtcc atttctgcgg catccagcca ggatacccgt cctcgctgac
     7981 gtaatatccc agcgccgcac cgctgtcatt aatctgcaca ccggcacggc agttccggct
     8041 gtcgccggta ttgttcgggt tgctgatgcg cttcgggctg accatccgga actgtgtccg
     8101 gaaaagccgc gacgaactgg tatcccaggt ggcctgaacg aacagttcac cgttaaaggc
     8161 gtgcatggcc acaccttccc gaatcatcat ggtaaacgtg cgttttcgct caacgtcaat
     8221 gcagcagcag tcatcctcgg caaactcttt ccatgccgct tcaacctcgc gggaaaaggc
     8281 acgggcttct tcctccccga tgcccagata gcgccagctt gggcgatgac tgagccggaa
     8341 aaaagacccg acgatatgat cctgatgcag ctaga
//
