LOCUS       sequence-292666-        4653 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  In vitro transcription of dominant negative form of P53 mRNA for RNA
            transfection into mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4653)
  AUTHORS   Watanabe T, Yamazaki S, Yoneda N, Shinohara H, Tomioka I, Higuchi Y,
            Yagoto M, Ema M, Suemizu H, Kawai K, Sasaki E
  TITLE     Highly efficient induction of primate iPS cells by combining RNA 
            transfection and chemical compounds.
  JOURNAL   Genes Cells. 2019 Jul;24(7):473-484. doi: 10.1111/gtc.12702. Epub 
            2019 Jun 17.
  PUBMED    31099158
REFERENCE   2  (bases 1 to 4653)
  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: "10.1111/gtc.12702";
            journalName: "Genes Cells"; date: "2019-07"; volume: "24"; issue: 
            "7"; pages: "473-484"
COMMENT     Sequence Label: phBG-P53DD
FEATURES             Location/Qualifiers
     source          1..4653
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(752..770)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(791..807)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(791..807)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(804..826)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    815..831
                     /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)."
     promoter        complement(join(839..845,846..863,864..869))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    884..905
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1022..1039)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1193..1781)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1273..1292)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             join(1905..1973,1974..2765)
                     /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(2123..2142)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     CDS             complement(3015..3809)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             complement(3015..3809)
                     /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
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3126..3145)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     3736..3755
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     CDS             complement(4158..4460)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
                     SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(4260..4279)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     primer_bind     4583..4605
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4597..4614
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4598..4614
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4621..4640
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4621..4639
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gcggccgctc ttctggtccc cacagactca gagagaaccc accaccatga ctgccatgga
       61 ggagtcacag tcggatatca gcctcaagag agcgctgccc acctgcacaa gcgcctctcc
      121 cccgcaaaag aaaaaaccac ttgatggaga gtatttcacc ctcaagatcc gcgggcgtaa
      181 acgcttcgag atgttccggg agctgaatga ggccttagag ttaaaggatg cccatgctac
      241 agaggagtct ggagacagca gggctcactc cagctacctg aagaccaaga agggccagtc
      301 tacttcccgc cataaaaaaa caatggtcaa gaaagtgggg cctgactcag actgactgcc
      361 tctgcatccc gtccccatca ccagcctccc cctctccttg ctgtcttatg acttcagggc
      421 tgagacacgg cgcgccagct cgctttcttg ctgtccaatt tctattaaag gttcctttgt
      481 tccctaagtc caactactaa actgggggat attatgaagg gccttgagca tctggattct
      541 gcctaataaa aaacatttat tttcattgct gcgtcgacag ctcgctttct tgctgtccaa
      601 tttctattaa aggttccttt gttccctaag tccaactact aaactggggg atattatgaa
      661 gggccttgag catctggatt ctgcctaata aaaaacattt attttcattg ctgcaagctt
      721 cacgaattcg tttaaacctg caggactagt ccctttagtg agggttaatt ctgagcttgg
      781 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
      841 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
      901 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
      961 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     1021 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     1081 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     1141 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     1201 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     1261 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     1321 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     1381 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     1441 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     1501 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     1561 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     1621 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     1681 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     1741 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     1801 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagac
     1861 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     1921 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     1981 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     2041 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     2101 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     2161 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     2221 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     2281 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     2341 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     2401 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     2461 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     2521 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     2581 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     2641 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     2701 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     2761 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt
     2821 aatttaaaag gatctaggtg aagatccttt ttgataatct catgagcgga tacatatttg
     2881 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     2941 ctgatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggaaattg
     3001 taagcgttaa taattcagaa gaactcgtca agaaggcgat agaaggcgat gcgctgcgaa
     3061 tcgggagcgg cgataccgta aagcacgagg aagcggtcag cccattcgcc gccaagctct
     3121 tcagcaatat cacgggtagc caacgctatg tcctgatagc ggtccgccac acccagccgg
     3181 ccacagtcga tgaatccaga aaagcggcca ttttccacca tgatattcgg caagcaggca
     3241 tcgccatggg tcacgacgag atcctcgccg tcgggcatgc tcgccttgag cctggcgaac
     3301 agttcggctg gcgcgagccc ctgatgctct tcgtccagat catcctgatc gacaagaccg
     3361 gcttccatcc gagtacgtgc tcgctcgatg cgatgtttcg cttggtggtc gaatgggcag
     3421 gtagccggat caagcgtatg cagccgccgc attgcatcag ccatgatgga tactttctcg
     3481 gcaggagcaa ggtgagatga caggagatcc tgccccggca cttcgcccaa tagcagccag
     3541 tcccttcccg cttcagtgac aacgtcgagc acagctgcgc aaggaacgcc cgtcgtggcc
     3601 agccacgata gccgcgctgc ctcgtcttgc agttcattca gggcaccgga caggtcggtc
     3661 ttgacaaaaa gaaccgggcg cccctgcgct gacagccgga acacggcggc atcagagcag
     3721 ccgattgtct gttgtgccca gtcatagccg aatagcctct ccacccaagc ggccggagaa
     3781 cctgcgtgca atccatcttg ttcaatcatg cgaaacgatc ctcatcctgt ctcttgatca
     3841 gagcttgatc ccctgcgcca tcagatcctt ggcggcaaga aagccatcca gtttactttg
     3901 cagggcttcc caaccttacc agagggcgcc ccagctggca attccggttc gcttgctgtc
     3961 cataaaaccg cccagtctag ctatcgccat gtaagcccac tgcaagctac ctgctttctc
     4021 tttgcgcttg cgttttccct tgtccagata gcccagtagc tgacattcat ccggggtcag
     4081 caccgtttct gcggactggc tttctacgtg aaaaggatct aggtgaagat cctttttgat
     4141 aatctcatgc ctgacattta tattccccag aacatcaggt taatggcgtt tttgatgtca
     4201 ttttcgcggt ggctgagatc agccacttct tccccgataa cggagaccgg cacactggcc
     4261 atatcggtgg tcatcatgcg ccagctttca tccccgatat gcaccaccgg gtaaagttca
     4321 cgggagactt tatctgacag cagacgtgca ctggccaggg ggatcaccat ccgtcgcccc
     4381 ggcgtgtcaa taatatcact ctgtacatcc acaaacagac gataacggct ctctctttta
     4441 taggtgtaaa ccttaaactg ccgtacgtat aggctgcgca actgttggga agggcgatcg
     4501 gtgcgggcct cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcgatta
     4561 agttgggtaa cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagtgaattg
     4621 taatacgact cactataggg cgaattgaat tta
//
