LOCUS       Exported                4653 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  In vitro transcription of dominant negative form of P53 mRNA for RNA
            transfection into mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    phBG-P53DD
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 May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4653
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(785..803)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(824..840)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(824..840)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(837..859)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    848..864
                     /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(872..902)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    917..938
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1055..1072)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1226..1814)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1306..1325)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             1938..2798
                     /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(2156..2175)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     CDS             complement(3048..3842)
                     /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(3159..3178)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     3769..3788
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     CDS             complement(4191..4493)
                     /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(4293..4312)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     primer_bind     4616..4638
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4630..4647
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4631..4647
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 taatacgact cactataggg cgaattgaat ttagcggccg ctcttctggt ccccacagac
       61 tcagagagaa cccaccacca tgactgccat ggaggagtca cagtcggata tcagcctcaa
      121 gagagcgctg cccacctgca caagcgcctc tcccccgcaa aagaaaaaac cacttgatgg
      181 agagtatttc accctcaaga tccgcgggcg taaacgcttc gagatgttcc gggagctgaa
      241 tgaggcctta gagttaaagg atgcccatgc tacagaggag tctggagaca gcagggctca
      301 ctccagctac ctgaagacca agaagggcca gtctacttcc cgccataaaa aaacaatggt
      361 caagaaagtg gggcctgact cagactgact gcctctgcat cccgtcccca tcaccagcct
      421 ccccctctcc ttgctgtctt atgacttcag ggctgagaca cggcgcgcca gctcgctttc
      481 ttgctgtcca atttctatta aaggttcctt tgttccctaa gtccaactac taaactgggg
      541 gatattatga agggccttga gcatctggat tctgcctaat aaaaaacatt tattttcatt
      601 gctgcgtcga cagctcgctt tcttgctgtc caatttctat taaaggttcc tttgttccct
      661 aagtccaact actaaactgg gggatattat gaagggcctt gagcatctgg attctgccta
      721 ataaaaaaca tttattttca ttgctgcaag cttcacgaat tcgtttaaac ctgcaggact
      781 agtcccttta gtgagggtta attctgagct tggcgtaatc atggtcatag ctgtttcctg
      841 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
      901 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
      961 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     1021 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     1081 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     1141 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     1201 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     1261 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     1321 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     1381 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     1441 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     1501 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     1561 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     1621 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     1681 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     1741 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     1801 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     1861 aaaactcacg ttaagggatt ttggtcatga gacaataacc ctgataaatg cttcaataat
     1921 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg
     1981 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg
     2041 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc
     2101 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat
     2161 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact
     2221 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca
     2281 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
     2341 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg
     2401 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg
     2461 agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg
     2521 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg
     2581 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag
     2641 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc
     2701 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga
     2761 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat
     2821 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     2881 tttttgataa tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     2941 gggttccgcg cacatttccc cgaaaagtgc cacctgatgc ggtgtgaaat accgcacaga
     3001 tgcgtaagga gaaaataccg catcaggaaa ttgtaagcgt taataattca gaagaactcg
     3061 tcaagaaggc gatagaaggc gatgcgctgc gaatcgggag cggcgatacc gtaaagcacg
     3121 aggaagcggt cagcccattc gccgccaagc tcttcagcaa tatcacgggt agccaacgct
     3181 atgtcctgat agcggtccgc cacacccagc cggccacagt cgatgaatcc agaaaagcgg
     3241 ccattttcca ccatgatatt cggcaagcag gcatcgccat gggtcacgac gagatcctcg
     3301 ccgtcgggca tgctcgcctt gagcctggcg aacagttcgg ctggcgcgag cccctgatgc
     3361 tcttcgtcca gatcatcctg atcgacaaga ccggcttcca tccgagtacg tgctcgctcg
     3421 atgcgatgtt tcgcttggtg gtcgaatggg caggtagccg gatcaagcgt atgcagccgc
     3481 cgcattgcat cagccatgat ggatactttc tcggcaggag caaggtgaga tgacaggaga
     3541 tcctgccccg gcacttcgcc caatagcagc cagtcccttc ccgcttcagt gacaacgtcg
     3601 agcacagctg cgcaaggaac gcccgtcgtg gccagccacg atagccgcgc tgcctcgtct
     3661 tgcagttcat tcagggcacc ggacaggtcg gtcttgacaa aaagaaccgg gcgcccctgc
     3721 gctgacagcc ggaacacggc ggcatcagag cagccgattg tctgttgtgc ccagtcatag
     3781 ccgaatagcc tctccaccca agcggccgga gaacctgcgt gcaatccatc ttgttcaatc
     3841 atgcgaaacg atcctcatcc tgtctcttga tcagagcttg atcccctgcg ccatcagatc
     3901 cttggcggca agaaagccat ccagtttact ttgcagggct tcccaacctt accagagggc
     3961 gccccagctg gcaattccgg ttcgcttgct gtccataaaa ccgcccagtc tagctatcgc
     4021 catgtaagcc cactgcaagc tacctgcttt ctctttgcgc ttgcgttttc ccttgtccag
     4081 atagcccagt agctgacatt catccggggt cagcaccgtt tctgcggact ggctttctac
     4141 gtgaaaagga tctaggtgaa gatccttttt gataatctca tgcctgacat ttatattccc
     4201 cagaacatca ggttaatggc gtttttgatg tcattttcgc ggtggctgag atcagccact
     4261 tcttccccga taacggagac cggcacactg gccatatcgg tggtcatcat gcgccagctt
     4321 tcatccccga tatgcaccac cgggtaaagt tcacgggaga ctttatctga cagcagacgt
     4381 gcactggcca gggggatcac catccgtcgc cccggcgtgt caataatatc actctgtaca
     4441 tccacaaaca gacgataacg gctctctctt ttataggtgt aaaccttaaa ctgccgtacg
     4501 tataggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag
     4561 ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag
     4621 tcacgacgtt gtaaaacgac ggccagtgaa ttg
//
