LOCUS       sequence-256372-        7748 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 7748)
  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 7748)
  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..7748
                     /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   4305..4367
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(4497..4514)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4497..4513)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4506..4528)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4655..5110
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4742..4761)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4952..4973
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5136..5240
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5241..5309,5310..6101)
                     /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(5459..5478)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6272..6860
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6761..6780
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7014..7031
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7148..7169
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7184..7189,7190..7207,7208..7214)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7222..7238
                     /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     7227..7249
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7246..7262
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7246..7262
                     /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 gccataccaa tggagctcac ggggacagcc
     3901 cccccccaaa gcccccaggg atgtaattac gtccctcccc cgctaggggg cagcagcgag
     3961 ccgcccgggg ctccgctccg gtccggcgct ccccccgcat ccccgagccg gcagcgtgcg
     4021 gggacagccc gggcacgggg aaggtggcac gggatcgctt tcctctgaac gcttctcgct
     4081 gctctttgag cctgcagaca cctgggggga tacggggaaa aagaattcga taaaagtttt
     4141 gttactttat agaagaaatt ttgagttttt gtttttttta ataaataaat aaacataaat
     4201 aaattgtttg ttgaatttat tattagtatg taagtgtaaa tataataaaa cttaatatct
     4261 attcaaatta ataaataaac ctcgatatac agaccgataa aacacatgcg tcaattttac
     4321 acatgattat ctttaacgta cgtcacaata tgattatctt tctagggtta atctagctgc
     4381 gtgttctgca gcgtgtcgag catcttcatc tgctccatca cgctgtaaaa cacatttgca
     4441 ccgcgagtct gcccgtcctc cacgggttca aaaacgtgaa tgaacgaggc gcgctcactg
     4501 gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt
     4561 gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct
     4621 tcccaacagt tgcgcagcct gaatggcgaa tgggacgcgc cctgtagcgg cgcattaagc
     4681 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     4741 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     4801 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     4861 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     4921 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     4981 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     5041 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     5101 cttacaattt aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     5161 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     5221 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     5281 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     5341 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     5401 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     5461 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     5521 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     5581 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     5641 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     5701 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     5761 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     5821 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     5881 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     5941 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     6001 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     6061 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     6121 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     6181 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     6241 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     6301 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     6361 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     6421 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     6481 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     6541 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     6601 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     6661 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     6721 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     6781 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     6841 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     6901 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     6961 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     7021 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     7081 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     7141 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc
     7201 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     7261 accatgatta cgccaagcgc gcccgccggg taactcacgg ggtatccatg tccatttctg
     7321 cggcatccag ccaggatacc cgtcctcgct gacgtaatat cccagcgccg caccgctgtc
     7381 attaatctgc acaccggcac ggcagttccg gctgtcgccg gtattgttcg ggttgctgat
     7441 gcgcttcggg ctgaccatcc ggaactgtgt ccggaaaagc cgcgacgaac tggtatccca
     7501 ggtggcctga acgaacagtt caccgttaaa ggcgtgcatg gccacacctt cccgaatcat
     7561 catggtaaac gtgcgttttc gctcaacgtc aatgcagcag cagtcatcct cggcaaactc
     7621 tttccatgcc gcttcaacct cgcgggaaaa ggcacgggct tcttcctccc cgatgcccag
     7681 atagcgccag cttgggcgat gactgagccg gaaaaaagac ccgacgatat gatcctgatg
     7741 cagctaga
//
