LOCUS       Exported               13592 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pTT-PB-SOKMLN-Cer, complete sequence.
ACCESSION   KM279352
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13592)
  AUTHORS   Debowski K, Warthemann R, Lentes J, Salinas-Riester G, Dressel R, 
            Langenstroth D, Gromoll J, Sasaki E, Behr R.
  TITLE     Non-Viral Generation of Marmoset Monkey iPS Cells by a 
            Six-Factor-in-One-Vector Approach
  JOURNAL   PLoS ONE 10 (3), E0118424 (2015)
  PUBMED    25785453
REFERENCE   2  (bases 1 to 13592)
  AUTHORS   Debowski K.
  TITLE     Direct Submission
  JOURNAL   Submitted (11-AUG-2014) Stem Cell Biology Unit, German Primate 
            Center, Kellnerweg 4, Goettingen, Niedersachsen 37077, Germany
REFERENCE   3  (bases 1 to 13592)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..13592
                     /organism="Cloning vector pTT-PB-SOKMLN-Cer"
                     /mol_type="other DNA"
                     /label=derived from Callithrix jacchus
                     /note="derived from Callithrix jacchus"
                     /db_xref="taxon:1633624"
     promoter        4..318
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    12..30
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    48..66
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    83..101
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    119..137
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    155..173
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    190..208
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    226..244
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     repeat_region   345..871
                     /label=piggyBac 5 terminal repeat
                     /note="piggyBac 5 terminal repeat"
     regulatory      878..2617
                     /regulatory_class="promoter"
                     /note="CAG promoter"
     enhancer        904..1283
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1286..1561
                     /label=chicken beta-actin promoter
     intron          1562..2573
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     misc_feature    2696..10913
                     /note="multiprotein reprogramming cassette; cDNA encoding 
                     one single mRNA with one ORF for the six reprogramming 
                     factors SOX2, OCT4, KLF4, cMYC, LIN28 and NANOG subdivided 
                     by 2A peptides followed by an IRES and the ORF for the 
                     fluorescent protein Cerulean"
     CDS             2696..9316
                     /codon_start=1
                     /product="fusion protein"
                     /label=fusion protein
                     /note="contains reprogramming factors SOX2, OCT4, KLF4, 
                     cMYC, LIN28, NANOG; ribosomal skip after amino acids 340, 
                     720, 1210, 1671 and 1900"
                     /protein_id="AJZ68903.1"
                     /translation="MYNMMETELKPPGPQQTSGGGGGNPTAAAAAAGGNQKNSPDRVKR
                     PMNAFMVWSRGQRRKMAQENPKMHNSEISKRLGAEWKLLSETEKRPFIDEAKRLRALHM
                     KEHPDYKYRPRRKTKTLMKKDKYTLPGGLLAPGGNGMAGGVGVGAGLGAGVNQRMDSYA
                     HMNGWSNGSYSMMQEQLGYPQHPGLNAHGAAQMQPMHRYDVSALQYNSMTSSQTYMNGS
                     PTYSMSYSQQGTPGMALGSMGSVVKSEASSSPPVVTSSSHSRAPCQAGDLRDMISMYLP
                     GAEVPEPAAPSRLHMSQHYQSGPVPGTAINGTLPLSHMVKQTLNFDLLKLAGDVESNPG
                     PGSMAGHLASDFAFSPPPGGGGDGPGGPETGWVDPRTWLSFQGPPGGPGIGPGVGPGAE
                     VWGIPPCPPPYEFCGGMAYCGPQVGVGLVPQGGLETSQPEGEAGAGVESNSEGASPEPC
                     TIPPGAVKLEKEKLEQNTEESQDIKALQKELEQFAKLLKQKRITLGYTQADVGLTLGVL
                     FGKVFSQTTICRFEALQLSFKNMCKLRPLLQKWVEEADNNENLQEICKAETLVQARKRK
                     RTSIENRVRGSLENLFLQCPKPTLQQISHIAQQLGLEKDVVRVWFCNRRQKGKRSSSDY
                     AQREDFEAAGSPFSGGPVSFPLAPGPHFGTPGYGSPHFTALYSSVPFPEGEAFPPVSVP
                     TLGSPMHSNEGRGSLLTCGDVEENPGPTRMAVSDALLPSFSTFASGPAGREKTLRQAGA
                     PNNRWREELSHMKRLPQVLPGRPYDLAAATVATDLESGGAGAACGGSNLAPLPRRETEE
                     FNDLLDLDFILSNSLSHPESVAATVSSSASASSSSSPTSSGPASAPSTCSFSYPIRAGS
                     DPGVAPGSTGGGLLYGRESAPPPTAPFNLADINDVSPSGGFVAELLRPELDPVYIPPQQ
                     PQPPGGGLMGKFVLKASLPPGSEYGSPSVISVSKGSPDGSHPVVVAPYNGGPPRMCPKI
                     KQEAVSSCTHLGAGPPLSNGHRPAAHDFPLGRQLPSRTTPTLGPEELLSSRDCHPALPL
                     PPGFHPHPGPNYPSFLPDQMQPQVPPLHYQELMPPGSCMPEEPKPKRGRRSWPRKRTAT
                     HTCDYAGCGKTYTKSSHLKAHLRTHTGEKPYHCDWDGCGWKFARSDELTRHYRKHTGHR
                     PFQCQKCDRAFSRSDHLALHMKRHFQCTNYALLKLAGDVESNPGPAAAMPLNVSFSNRN
                     YDLDYDSVQPYFYCDEEENFYQQQQQSELQPPAPSEDIWKKFELLPTPPLSPSRRSGLC
                     STSCVSVTPFSPRGDNDGGGGSFSTADQLEMVTELLGGDMVNQSFICDPDDETFIKNII
                     IQDCMWSGFSAAAKLVSEKLASYQAARKDSSSPNPARGHSVCSTSSLYLQDLSAAASEC
                     IDPSVVFPYPLNDSSSPKPCASPDSSAFSTSSDSLLSSTESSPRASPEPLVLHEETPPT
                     TSSDSEEEQEDEEIDVVSVEKRQPPGKRSESGSPSSGGHSKPPHSPLVLKRCHVSTHQH
                     NYAAPPSTRKDYPAAKRVKLDSVRVLRQISNNRKCTSPRSSDTEENDKRRTHNVLERQR
                     RNELKRSFFALRDQIPELENNEKAPKVVILKKATTYILSVQAEEQKLISEKDLLRKRRE
                     QLKHKLEQLRNSCATGEGRGSLLTCGDVEENPGPMGSVSNQQFAGGCAKAAEEAPEEAP
                     EDAARAADEPQLLHGAGICKWFNVRMGFGFLSMTARAGVALDPPVDVFVHQSKLHMEGF
                     RSLKEGEAVEFTFKKSAKGLESIRVTGPGGVFCIGSERRPKGKNMQKRRSKGDRCYNCG
                     GLDHHAKECKLPPQPKKCHFCQSISHMVASCPLKAQQAPSAQGKPTYFREEEEEIHSPT
                     LLPEAQNFEEGRGSLLTCGDVEENPGPMSMDPVCSQSLPCSEASDCGESSPTPVICEPE
                     ENYPSLQMCSAEMPHTETVSPLPSSMDLLTQDSPDSSTSPKGKRPTAAESSPAVKEDKV
                     PVKKQKTRTVFSSTQLCVLNDRFQRQKYLSLQQMQELSNILNLSYKQVKTWFQNQRMKS
                     KRWQKNNWPRNTSDVTQKASAPTYPSLYSSYHQGCLVNTAGNLPVWNNQTWSNSTWSNQ
                     TRNIQSWSNHSWNTQTWCTQSWNTQAWSSPFYNCGEEPLQSCMQFQPNSPASDLEAALE
                     TAGENHNVIQQTTRYLNTSQTMDLFLNYSTNMPPEDV"
     CDS             3653..3718
                     /codon_start=1
                     /product="2A peptide from foot-and-mouth disease virus 
                     polyprotein"
                     /label=F2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="VKQTLNFDLLKLAGDVESNPGP"
     CDS             4805..4858
                     /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             6269..6328
                     /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             7292..7318
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             7562..7591
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEKDL"
     CDS             7658..7711
                     /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             8345..8398
                     /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"
     regulatory      9341..9926
                     /regulatory_class="ribosome_binding_site"
                     /note="IRES, internal ribosomal entry site"
     misc_feature    9345..9912
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     CDS             9938..10657
                     /codon_start=1
                     /product="fluorescent protein cerulean"
                     /label=fluorescent protein cerulean
                     /protein_id="AJZ68902.1"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     polyA_signal    10689..10913
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   10939..11388
                     /label=piggyBac 3 terminal repeat
                     /note="piggyBac 3 terminal repeat"
     polyA_signal    11506..11587
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(11769..12357)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(12528..13388)
                     /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"
     promoter        complement(13389..13493)
                     /gene="bla"
                     /label=AmpR promoter