LOCUS       sequence-268039-        7749 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 7749)
  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 7749)
  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-ChiNW
FEATURES             Location/Qualifiers
     source          1..7749
                     /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(1307..1881,1882..1884,1885..1893)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1474..1491)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     1701..1720
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             1906..2700
                     /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(1960..1979)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2570..2589
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     misc_feature    2725..3313
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2778..2798)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3196..3207)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3393..3527
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3417..3436)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3471..3490
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4306..4368
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(4498..4515)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4498..4514)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4507..4529)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4656..5111
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4743..4762)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4953..4974
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5137..5241
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5242..5310,5311..6102)
                     /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(5460..5479)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6273..6861
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6762..6781
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7015..7032
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7149..7170
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7185..7190,7191..7208,7209..7215)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7223..7239
                     /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     7228..7250
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7247..7263
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7247..7263
                     /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 tagcgaattc gaatttaaat cggatccgcg gccgcgcccc tctccctccc
     1321 ccccccctaa cgttactggc cgaagccgct tggaataagg ccggtgtgcg tttgtctata
     1381 tgttattttc caccatattg ccgtcttttg gcaatgtgag ggcccggaaa cctggccctg
     1441 tcttcttgac gagcattcct aggggtcttt cccctctcgc caaaggaatg caaggtctgt
     1501 tgaatgtcgt gaaggaagca gttcctctgg aagcttcttg aagacaaaca acgtctgtag
     1561 cgaccctttg caggcagcgg aaccccccac ctggcgacag gtgcctctgc ggccaaaagc
     1621 cacgtgtata agatacacct gcaaaggcgg cacaacccca gtgccacgtt gtgagttgga
     1681 tagttgtgga aagagtcaaa tggctctcct caagcgtatt caacaagggg ctgaaggatg
     1741 cccagaaggt accccattgt atgggatctg atctggggcc tcggtgcaca tgctttacat
     1801 gtgtttagtc gaggttaaaa aaacgtctag gccccccgaa ccacggggac gtggttttcc
     1861 tttgaaaaac acgatgataa tatggccaca accatggcgt ccggaatgat tgaacaagat
     1921 ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca
     1981 caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg
     2041 gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg
     2101 cggctatcgt ggctggccgc gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact
     2161 gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct
     2221 caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg
     2281 cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt
     2341 actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc
     2401 gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc
     2461 gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga
     2521 ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc
     2581 cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt
     2641 atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga
     2701 gtcgctagag ttcgcgtctg gaacaatcaa cctctggatt acaaaatttg tgaaagattg
     2761 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct
     2821 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg
     2881 ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact
     2941 gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc
     3001 gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc
     3061 cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaag
     3121 ctgacgtcct ttccatggct gctcgcctgt gttgccacct ggattctgcg cgggacgtcc
     3181 ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg
     3241 gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg
     3301 gccgcctccc cgcctggaat taattctggc taagcacttc gtggccgtcg atcgtttaaa
     3361 gggaggtagt gagtcgagat atcgatacta gtgatccaga catgataaga tacattgatg
     3421 agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg
     3481 atgctattgc tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt
     3541 gcattcattt tatgtttcag gttcaggggg aggtgtggga ggttttttaa agcaagtaaa
     3601 acctctacaa atgtggtatg gctgattatg atcctctaga gggacagccc ccccccaaag
     3661 cccccaggga tgtaattacg tccctccccc gctagggggc agcagcgagc cgcccggggc
     3721 tccgctccgg tccggcgctc cccccgcatc cccgagccgg cagcgtgcgg ggacagcccg
     3781 ggcacgggga aggtggcacg ggatcgcttt cctctgaacg cttctcgctg ctctttgagc
     3841 ctgcagacac ctggggggat acggggaaaa agccatacca atggagctca cggggacagc
     3901 ccccccccaa agcccccagg gatgtaatta cgtccctccc ccgctagggg gcagcagcga
     3961 gccgcccggg gctccgctcc ggtccggcgc tccccccgca tccccgagcc ggcagcgtgc
     4021 ggggacagcc cgggcacggg gaaggtggca cgggatcgct ttcctctgaa cgcttctcgc
     4081 tgctctttga gcctgcagac acctgggggg atacggggaa aaagaattcg ataaaagttt
     4141 tgttacttta tagaagaaat tttgagtttt tgtttttttt aataaataaa taaacataaa
     4201 taaattgttt gttgaattta ttattagtat gtaagtgtaa atataataaa acttaatatc
     4261 tattcaaatt aataaataaa cctcgatata cagaccgata aaacacatgc gtcaatttta
     4321 cacatgatta tctttaacgt acgtcacaat atgattatct ttctagggtt aatctagctg
     4381 cgtgttctgc agcgtgtcga gcatcttcat ctgctccatc acgctgtaaa acacatttgc
     4441 accgcgagtc tgcccgtcct ccacgggttc aaaaacgtga atgaacgagg cgcgctcact
     4501 ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct
     4561 tgcagcacat ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc
     4621 ttcccaacag ttgcgcagcc tgaatggcga atgggacgcg ccctgtagcg gcgcattaag
     4681 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     4741 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     4801 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     4861 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     4921 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     4981 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
     5041 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac
     5101 gcttacaatt taggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
     5161 ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa
     5221 taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt
     5281 tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat
     5341 gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag
     5401 atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg
     5461 ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata
     5521 cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat
     5581 ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc
     5641 aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg
     5701 ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac
     5761 gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact
     5821 ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa
     5881 gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct
     5941 ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc
     6001 tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga
     6061 cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac
     6121 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     6181 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     6241 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     6301 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     6361 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
     6421 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     6481 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     6541 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     6601 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     6661 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     6721 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     6781 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     6841 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     6901 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     6961 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
     7021 tcagtgagcg aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg
     7081 ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc
     7141 aacgcaatta atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt
     7201 ccggctcgta tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat
     7261 gaccatgatt acgccaagcg cgcccgccgg gtaactcacg gggtatccat gtccatttct
     7321 gcggcatcca gccaggatac ccgtcctcgc tgacgtaata tcccagcgcc gcaccgctgt
     7381 cattaatctg cacaccggca cggcagttcc ggctgtcgcc ggtattgttc gggttgctga
     7441 tgcgcttcgg gctgaccatc cggaactgtg tccggaaaag ccgcgacgaa ctggtatccc
     7501 aggtggcctg aacgaacagt tcaccgttaa aggcgtgcat ggccacacct tcccgaatca
     7561 tcatggtaaa cgtgcgtttt cgctcaacgt caatgcagca gcagtcatcc tcggcaaact
     7621 ctttccatgc cgcttcaacc tcgcgggaaa aggcacgggc ttcttcctcc ccgatgccca
     7681 gatagcgcca gcttgggcga tgactgagcc ggaaaaaaga cccgacgata tgatcctgat
     7741 gcagctaga
//
