LOCUS       Exported                9414 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  To express Oct4, Klf4, Sox2, c-Myc and hairpin RNA p53. A single 
            plasmid reprogramming system using a mini-intronic plasmid (MIP)..
ACCESSION   .
VERSION     .
KEYWORDS    MIP 247 CoMiP 4in1 with shRNA p53
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9414)
  AUTHORS   Diecke S, Lu J, Lee J, Termglinchan V, Kooreman NG, Burridge PW, 
            Ebert AD, Churko JM, Sharma A, Kay MA, Wu JC
  TITLE     Novel codon-optimized mini-intronic plasmid for efficient, 
            inexpensive, and xeno-free induction of pluripotency.
  JOURNAL   Sci Rep. 2015 Jan 28;5:8081. doi: 10.1038/srep08081.
  PUBMED    25628230
REFERENCE   2  (bases 1 to 9414)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..9414
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        46..452
                     /label=SFFV promoter
                     /note="spleen focus-forming virus long terminal repeat 
                     (LTR) promoter"
     primer_bind     complement(578..595)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(749..1337)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(829..848)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     regulatory      1827..1836
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             2922..2978
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             4401..4454
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             5415..5474
                     /codon_start=1
                     /product="2A peptide from equine rhinitis A virus 
                     polyprotein"
                     /label=E2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="QCTNYALLKLAGDVESNPGP"
     CDS             5478..6797
                     /codon_start=1
                     /product="human c-Myc proto-oncogene"
                     /label=c-Myc
                     /translation="MPLNVSFTNRNYDLDYDSVQPYFYCDEEENFYQQQQQSELQPPAP
                     SEDIWKKFELLPTPPLSPSRRSGLCSPSYVAVTPFSLRGDNDGGGGSFSTADQLEMVTE
                     LLGGDMVNQSFICDPDDETFIKNIIIQDCMWSGFSAAAKLVSEKLASYQAARKDSGSPN
                     PARGHSVCSTSSLYLQDLSAAASECIDPSVVFPYPLNDSSSPKSCASQDSSAFSPSSDS
                     LLSSTESSPQGSPEPLVLHEETPPTTSSDSEEEQEDEEEIDVVSVEKRQAPGKRSESGS
                     PSAGGHSKPPHSPLVLKRCHVSTHQHNYAAPPSTRKDYPAAKRVKLDSVRVLRQISNNR
                     KCTSPRSSDTEENVKRRTHNVLERQRRNELKRSFFALRDQIPELENNEKAPKVVILKKA
                     TAYILSVQAEEQKLISEEDLLRKRREQLKHKLEQLRNSCA"
     misc_feature    6832..7418
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(6999..7016)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     7226..7245
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             7419..8120
                     /codon_start=1
                     /product="dimeric variant of DsRed fluorescent protein 
                     (Shaner et al., 2004)"
                     /label=dTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLYGMDELYK"
     primer_bind     complement(7552..7570)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     7820..7839
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     8014..8037
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     misc_feature    8139..8727
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8192..8212)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8610..8621)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    8739..8963
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(9121..9141)
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
ORIGIN
        1 ggctccccgg gcgcgactag tgaattcaga tctgagctag ctgcagtaac gccattttgc
       61 aaggcatgga aaaataccaa accaagaata gagaagttca gatcaagggc gggtacatga
      121 aaatagctaa cgttgggcca aacaggatat ctgcggtgag cagtttcggc cccggcccgg
      181 ggccaagaac agatggtcac cgcagtttcg gccccggccc gaggccaaga acagatggtc
      241 cccagatatg gcccaaccct cagcagtttc ttaagaccca tcagatgttt ccaggctccc
      301 ccaaggacct gaaatgaccc tgcgccttat ttgaattaac caatcagcct gcttctcgct
      361 tctgttcgcg cgcttctgct tcccgagctc tataaaagag ctcacaaccc ctcactcggc
      421 gcgccagtcc tccgacagac tgagtcggcc ggtggatcca ccggtattgg gatcttcaca
      481 cagcaggtaa ggttgcgggc cgggcctggg ccgggtccgg gccgggtatt gcccgcctaa
      541 tgagcgggct tttttttctt accccttctt ccgcttcctc gctcactgac tcgctgcgct
      601 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
      661 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
      721 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
      781 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
      841 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
      901 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
      961 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     1021 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     1081 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     1141 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     1201 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     1261 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     1321 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     1381 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     1441 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     1501 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     1561 catccatagt tgcctgactc ctgcaaacca cgttgtggta gaattggtaa agagagtcgt
     1621 gtaaaatatc gagttcgcac atcttgttgt ctgattattg atttttggcg aaaccatttg
     1681 atcatatgac aagatgtgta tctaccttaa cttaatgatt ttgataaaaa tcattaggta
     1741 ccccggcccg cactgacccc tggtgttgct tttttttttt aggccgcaag ctgaagcgtg
     1801 tccactagct gactagctaa accggtgcca ccatggccgg acacctggcc agcgacttcg
     1861 cctttagccc acctcctggc ggagggggag atggacctgg cggccctgaa cctggctggg
     1921 tggaccctcg gacctggctg tcctttcaag gccctccagg cggacctgga attggaccag
     1981 gcgtgggccc tggcagcgaa gtgtggggca tccctccctg ccctcccccc tacgagtttt
     2041 gcggcggcat ggcctactgt ggacctcagg tcggcgtggg actggtgcct cagggcggcc
     2101 tggaaacctc tcagcctgag ggcgaagccg gcgtgggcgt ggagagcaac tctgacggcg
     2161 ccagccctga gccttgtacc gtgaccccag gcgccgtgaa gctggaaaaa gagaagctgg
     2221 aacagaaccc cgaggaaagc caggacatca aggccctgca gaaagaactg gaacagttcg
     2281 ccaagctgct gaagcagaag cggatcaccc tgggctacac acaggctgac gtgggactga
     2341 cactgggcgt gctgttcggc aaggtgttca gccagaccac catctgcaga ttcgaagccc
     2401 tgcagctgtc cttcaagaac atgtgcaagc tgcggcccct gctgcagaaa tgggtggagg
     2461 aagccgacaa caacgagaac ctgcaggaaa tctgcaaggc cgagacactg gtgcaggccc
     2521 ggaagcggaa gcggaccagc atcgagaaca gagtgcgggg caacctggaa aacctgttcc
     2581 tgcagtgccc caagcccacc ctgcagcaga tcagccacat tgcacagcag ctgggactgg
     2641 aaaaggacgt cgtgagagtg tggttctgca accggcggca gaagggcaag agaagcagca
     2701 gcgactacgc ccagagagag gacttcgagg ccgctggctc cccttttagc ggcggacccg
     2761 tgtcctttcc tctggcccct ggccctcact ttggcacccc tggctacggc agcccacact
     2821 tcaccgccct gtacagcagc gtgccctttc ccgagggcga ggccttccct cctgtgtccg
     2881 tgaccaccct gggcagcccc atgcatagca actcttccgg agccaccaac ttctccctgc
     2941 tgaagcaggc cggcgacgtg gaggagaacc ccggcccctc catggctgtg tccgacgccc
     3001 tgctgcccag cttcagcacc tttgcctccg gccctgctgg cagagagaaa acactgcggc
     3061 aggctggcgc ccctaacaac agatggcggg aggaactgag ccacatgaag cggctgcctc
     3121 ccgtgctgcc cggcagacct tatgatctgg ccgctgccac cgtggccacc gatctggaat
     3181 ctggcggagc cggcgctgcc tgtggcggaa gcaacctggc ccccctgccc agacgggaga
     3241 cagaggaatt caacgacctg ctggacctgg acttcatcct gagcaacagc ctgacccacc
     3301 ctcctgagtc tgtggccgcc accgtgtcta gcagcgccag cgccagcagc agctctagcc
     3361 ctagcagcag cggacctgcc agcgccccca gcacctgcag cttcacctac cccatcagag
     3421 ccggcaatga ccctggcgtg gcccctggcg gaacaggagg aggactgctg tacggcagag
     3481 agtccgcccc tccccccacc gcccccttca acctggccga catcaacgat gtgtccccca
     3541 gcggcggctt tgtggccgag ctgctgcgac ctgagctgga ccccgtgtac atcccccctc
     3601 agcagccaca gcctcccggc ggaggactga tgggcaagtt cgtgctgaag gccagcctga
     3661 gcgcccctgg cagcgagtac ggcagcccca gcgtgatcag cgtgtccaag ggcagccctg
     3721 atggctctca ccctgtggtg gtggcccctt acaatggcgg ccctcccaga acctgcccca
     3781 agatcaagca ggaagccgtc agcagctgta cacacctggg agccggccct ccactgagca
     3841 atggccacag acctgccgcc cacgacttcc ctctgggaag gcagctccct agcagaacca
     3901 cccctaccct gggcctggaa gaggtgctgt ccagcagaga ctgccaccct gccctgcctc
     3961 tgccacctgg ctttcaccct caccctggcc ccaactaccc cagcttcctg cccgaccaga
     4021 tgcagcctca ggtgcccccc ctgcactacc aagaactgat gcccccaggc agctgcatgc
     4081 ccgaggaacc caagcccaag cggggcagaa gaagctggcc ccggaagaga accgccacac
     4141 acacctgcga ttacgccggc tgcggcaaga cctacaccaa gagcagccac ctgaaggccc
     4201 acctgagaac ccacaccggc gagaagccct accactgcga ctgggacggc tgtggctgga
     4261 agttcgccag aagcgacgag ctgacccggc actacagaaa gcacaccggc caccggccct
     4321 tccagtgcca gaagtgcgac cgggccttca gcagaagcga ccacctggct ctgcacatga
     4381 agagacactt cggctccgga gagggccggg gctctctgct gacctgtggc gacgtggagg
     4441 agaaccccgg cccctccatg tacaacatga tggaaaccga gctgaagccc cctggccctc
     4501 agcaaacatc tggcggcgga gggggcaatt ctacagccgc cgctgccggc ggaaaccaga
     4561 agaacagccc cgacagagtg aagcggccca tgaacgcctt catggtctgg tcccgcggcc
     4621 agcggcggaa gatggcccaa gaaaacccca agatgcacaa cagcgagatc agcaagagac
     4681 tgggcgccga gtggaagctg ctgtccgaga cagagaagag acccttcatc gacgaggcca
     4741 agcggctgag agccctgcac atgaaagagc accccgacta caagtaccgg cccagaagaa
     4801 agaccaagac cctgatgaag aaggacaagt acaccctgcc tggcggactg ctggcccctg
     4861 gcggcaattc tatggccagc ggagtgggag tgggagctgg actgggagcc ggcgtgaacc
     4921 agcggatgga cagctacgcc cacatgaacg gctggtccaa cggcagctac agcatgatgc
     4981 aggaccagct gggctaccct cagcaccctg gcctgaatgc ccatggcgcc gctcagatgc
     5041 agcccatgca cagatacgac gtgtccgccc tgcagtacaa cagcatgacc agcagccaga
     5101 cctacatgaa tggcagcccc acctacagca tgtcctacag ccagcagggc acacctggca
     5161 tggccctggg cagcatgggc agcgtggtca agagcgaggc cagcagcagc cctcctgtgg
     5221 tcaccagcag ctcccacagc agagcccctt gtcaggccgg cgacctgcgg gacatgatca
     5281 gcatgtacct gccaggcgcc gaagtgcctg aacctgccgc ccctagcaga ctgcacatga
     5341 gccagcacta ccagagcggc cctgtgcctg gcaccgccat caatggcacc ctgcccctga
     5401 gccacatgac gcgtcagtgc acaaactacg ccctgttgaa gctggctgga gacgtggagt
     5461 ccaaccctgg accctccatg cccctcaacg ttagcttcac caacaggaac tatgacctcg
     5521 actacgactc ggtgcagccg tatttctact gcgacgagga ggagaacttc taccagcagc
     5581 agcagcagag cgagctgcag cccccggcgc ccagcgagga tatctggaag aaattcgagc
     5641 tgctgcccac cccgcccctg tcccctagcc gccgctccgg gctctgctcg ccctcctacg
     5701 ttgcggtcac acccttctcc cttcggggag acaacgacgg cggtggcggg agcttctcca
     5761 cggccgacca gctggagatg gtgaccgagc tgctgggagg agacatggtg aaccagagtt
     5821 tcatctgcga cccggacgac gagaccttca tcaaaaacat catcatccag gactgtatgt
     5881 ggagcggctt ctcggccgcc gccaagctcg tctcagagaa gctggcctcc taccaggctg
     5941 cgcgcaaaga cagcggcagc ccgaaccccg cccgcggcca cagcgtctgc tccacctcca
     6001 gcttgtacct gcaggatctg agcgccgccg cctcagagtg catcgacccc tcggtggtct
     6061 tcccctaccc tctcaacgac agcagctcgc ccaagtcctg cgcctcgcaa gactccagcg
     6121 ccttctctcc gtcctcggat tctctgctct cctcgacgga gtcctccccg cagggcagcc
     6181 ccgagcccct ggtgctccat gaggagacac cgcccaccac cagcagcgac tctgaggagg
     6241 aacaagaaga tgaggaagaa atcgatgttg tttctgtgga aaagaggcag gctcctggca
     6301 aaaggtcaga gtctggatca ccttctgctg gaggccacag caaacctcct cacagcccac
     6361 tggtcctcaa gaggtgccac gtctccacac atcagcacaa ctacgcagcg cctccctcca
     6421 ctcggaagga ctatcctgct gccaagaggg tcaagttgga cagtgtcaga gtcctgagac
     6481 agatcagcaa caaccgaaaa tgcaccagcc ccaggtcctc ggacaccgag gagaatgtca
     6541 agaggcgaac acacaacgtc ttggagcgcc agaggaggaa cgagctaaaa cggagctttt
     6601 ttgccctgcg tgaccagatc ccggagttgg aaaacaatga aaaggccccc aaggtagtta
     6661 tccttaaaaa agccacagca tacatcctgt ccgtccaagc agaggagcaa aagctcattt
     6721 ctgaagagga cttgttgcgg aaacgacgag aacagttgaa acacaaactt gaacagctac
     6781 ggaactcttg tgcgtaagtc gacgaattcg gtaccgcggg cccgggatcc gcccctctcc
     6841 ctcccccccc cctaacgtta ctggccgaag ccgcttggaa taaggccggt gtgcgtttgt
     6901 ctatatgtta ttttccacca tattgccgtc ttttggcaat gtgagggccc ggaaacctgg
     6961 ccctgtcttc ttgacgagca ttcctagggg tctttcccct ctcgccaaag gaatgcaagg
     7021 tctgttgaat gtcgtgaagg aagcagttcc tctggaagct tcttgaagac aaacaacgtc
     7081 tgtagcgacc ctttgcaggc agcggaaccc cccacctggc gacaggtgcc tctgcggcca
     7141 aaagccacgt gtataagata cacctgcaaa ggcggcacaa ccccagtgcc acgttgtgag
     7201 ttggatagtt gtggaaagag tcaaatggct ctcctcaagc gtattcaaca aggggctgaa
     7261 ggatgcccag aaggtacccc attgtatggg atctgatctg gggcctcggt gcacatgctt
     7321 tacatgtgtt tagtcgaggt taaaaaaacg tctaggcccc ccgaaccacg gggacgtggt
     7381 tttcctttga aaaacacgat gataatatgg ccacaaccat ggtgagcaag ggcgaggagg
     7441 tcatcaaaga gttcatgcgc ttcaaggtgc gcatggaggg ctccatgaac ggccacgagt
     7501 tcgagatcga gggcgagggc gagggccgcc cctacgaggg cacccagacc gccaagctga
     7561 aggtgaccaa gggcggcccc ctgcccttcg cctgggacat cctgtccccc cagttcatgt
     7621 acggctccaa ggcgtacgtg aagcaccccg ccgacatccc cgattacaag aagctgtcct
     7681 tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggt ctggtgaccg
     7741 tgacccagga ctcctccctg caggacggca cgctgatcta caaggtgaag atgcgcggca
     7801 ccaacttccc ccccgacggc cccgtaatgc agaagaagac catgggctgg gaggcctcca
     7861 ccgagcgcct gtacccccgc gacggcgtgc tgaagggcga gatccaccag gccctgaagc
     7921 tgaaggacgg cggccactac ctggtggagt tcaagaccat ctacatggcc aagaagcccg
     7981 tgcaactgcc cggctactac tacgtggaca ccaagctgga catcacctcc cacaacgagg
     8041 actacaccat cgtggaacag tacgagcgct ccgagggccg ccaccacctg ttcctgtacg
     8101 gcatggacga gctgtacaag taaagcggcc gcgtcgacaa tcaacctctg gattacaaaa
     8161 tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg
     8221 ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct
     8281 tgtataaatc ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg
     8341 gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct
     8401 gtcagctcct ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg
     8461 ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg
     8521 tgttgtcggg gaagctgacg tcctttccat ggctgctcgc ctgtgttgcc acctggattc
     8581 tgcgcgggac gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc
     8641 gcggcctgct gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc
     8701 ggatctccct ttgggccgcc tccccgcctg gactcgagct gtgccttcta gttgccagcc
     8761 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt
     8821 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct
     8881 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc
     8941 tggggatgcg gtgggctcta tggagatctg atatctctag tcccgggctg caggaattcc
     9001 aaaaaagact ccagtggtaa tctactctct tgaagtagat taccactgga gtcggcccaa
     9061 acaaggcttt tctccaaggg atatttatag tctcaaaaca cacaattact ttacagttag
     9121 ggtgagtttc cttttgtgct gttttttaaa ataataattt agtatttgta tctcttatag
     9181 aaatccaagc ctatcatgta aaatgtagct agtattaaaa agaacagatt atctgtcttt
     9241 tatcgcacat taagcctcta tagttactag gaaatattat atgcaaatta accggggcag
     9301 gggagtagcc gagcttctcc cacaagtctg tgcgaggggg ccggcgcggg cctagagatg
     9361 gcggcgtcac tagagtcgac ccatgggggc ccgccccaac tggggtaacc tttg
//
