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ID   PTON       preliminary; circular DNA; SYN; 2846 BP.
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AC   M59367; M34008;
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DT   21-DEC-1990 (Rel. 5, Created)
DT   01-JUL-1995 (Rel. 12, Last updated, Version 1)
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DE   E. coli plasmid vector pTO-N - complete.
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KW   cloning vector.
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OS   Cloning vector
OC   Artificial sequences; Cloning vehicles.
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RN   [1]
RP   1-2846
RC   pJPNPLA2 from bovine pancreas PLA2
RC   M13mp18-propla2 from M13mp18 & pJPNPLA2
RC   pET-5a-propla2 from M13mp18-propla2 & pET-5a
RC   pET-ompA from pET-3a & pIN-III-ompA3
RC   pTO from pET-ompA
RC   pTO-propla2 from pTO & pJPNPLA2
RC   M13mp18-propla2m from M13mp18-propla2
RC   M13mp18-ompA from M13mp18 & pIN-III-ompA3
RC   M13mp18-ompAm from M13mp18-ompA
RC   pTO-A2M from M13mp18-propla2m & M13mp18-ompAm & pTO
RC   pTO-N from pTO-A2M
RA   Deng T., Noel J.P., Tsai M.D.;
RT   "A novel expression vector for high-level synthesis and secretion of
RT   foreign proteins in Escherichia coli: Overproduction of bovine
RT   pancreatic phospholipase A-2";
RL   Gene 93:229-234(1990).
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RN   [2]
RC   pMF3 from pI258 & mini-F
RC   pMF21 from F factor
RA   Manis J.J., Kline B.C.;
RT   "Restriction endonuclease mapping and mutagenesis of the F sex
RT   factor replication region";
RL   Mol. Gen. Genet. 152:175-182(1977).
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CC   old Promega vector.
CC   NM (pTO-N)
CC   CM (yes)
CC   NA (ds-DNA)
CC   TP (circular)
CC   ST ()
CC   TY (plasmid)
CC   SP ()
CC   HO (E.coli)
CC   CP (high)
CC   FN (expression)(secretion)
CC   SE ()
CC   PA (pTO)
CC   BR ()
CC   OF ()
CC   OR ()
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FH   Key             Location/Qualifiers
FH
FT   misc_feature    0..0
FT                   /note="1. bovine pancreas PLA2
FT                   -> pJPNPLA2
FT                   1. M13mp18 remove EcoRI-BamHI 21bp 6231..6252,
FT                   \ MCS/7228bp
FT                   2. pJPNPLA2 EcoRI-BamHI 400bp, proPLA2
FT                   -> M13mp18-propla2 7600bp
FT                   1. M13mp18-propla2 EcoRI 7600bp
FT                   mutagenesis, to make NheI
FT                   NheI-BamHI 400bp, proPLA2
FT                   2. pET-5a remove small NheI-BamHI, MCS
FT                   -> pET-5a-propla2
FT                   1. pET-3a XbaI-BamHI 4500bp, missing part of MCS
FT                   2. pIN-III-ompA3 XbaI-BamHI 99bp, ompA
FT                   -> pET-ompA
FT                   1. pET-ompA EcoRI-PvuII 3200bp
FT                   Klenow:Klenow
FT                   EcoRV-EcoRV 2800bp
FT                   Klenow:Klenow
FT                   -> pTO
FT                   1. pTO remove EcoRI-BamHI, MCS
FT                   2. pJPNPLA2 EcoRI-BamHI 400bp, proPLA2
FT                   -> pTO-propla2
FT                   1. M13mp18-propla2 7600bp
FT                   mutagenesis, to make Eco47III in proPLA2
FT                   -> M13mp18-propla2m 7600bp
FT                   1. M13mp18 remove XbaI-HindIII 24bp 6258..6282,
FT                   \ MCS/7225bp
FT                   2. pIN-III-ompA3 XbaI-HindIII 93bp, ompA
FT                   -> M13mp18-ompA 7318bp
FT                   1. M13mp18-ompA 7318bp
FT                   mutagenesis, to make Eco47III in ompA
FT                   -> M13mp18-ompAm 7318bp
FT                   1. M13mp18-propla2m Eco47III-BamHI 380bp, proPLA2
FT                   2. M13mp18-ompAm Eco47III-XbaI 86bp, ompA
FT                   3. pTO large XbaI-BamHI 2400bp
FT                   -> pTO-A2M 2850bp
FT                   1. pTO-A2M EcoRI 2850bp
FT                   2. NcoI-EcoRI linker 12bp ccatgggaattc
FT                   -> pTO-N 2846bp"
FT   promoter        21..39
FT                   /note="PRO"
FT   misc_feature    81..143
FT                   /note="E. coli OmpA signal sequence"
FT   rep_origin      0..0
FT                   /note="ORI E. coli pMB1 (ColE1 and pBR322)"
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SQ   Sequence 2846 BP; 704 A; 722 C; 740 G; 680 T; 0 other;
     agatctcgat cccgcgaaat taatacgact cactataggg agaccacaac ggtttccctc
     tagataacga ggcgcaaaaa atgaaaaaga cagctatcgc gattgcagtg gcactggctg
     gtttcgctac cgtagcgcag gccatgggaa ttccaagctt ggatccggct gctaacaaag
     cccgaaagga agctgagttg gctgctgcca ccgctgagca ataactagca taaccccttg
     gggcctctaa acgggtcttg aggggttttt tgctgaaagg aggaactata tccggataat
     tcttgaagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata
     atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     gttctgctat gtggcgcggt attatcccgt gttgacgccg ggcaagagca actcggtcgc
     cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     ccaaacgacg agcgtgacac cacgatgcct gcagcaatgg caacaacgtt gcgcaaacta
     ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
     agcgagtcag tgagcgagga agcggaagag cgcctgatgc ggtattttct ccttacgcat
     ctgtgcggta tttcacaccg catatggtgc actctcagta caatctgctc tgatgccgca
     tagttaagcc agtatacact ccgctatcgc tacgtgactg ggtcatggct gcgccccgac
     acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     aacgcgcgag gcagattagg aagcagccca gtagtaggtt gaggccgttg agcaccgccg
     ccgcaaggaa tggtgcatgc aaggagatgg cgcccaacag tcccccggcc acggggcctg
     ccaccatacc cacgccgaaa caagcgctca tgagcccgaa gtggcgagcc cgatcttccc
     catcggtgat gtcggcgata taggcgccag caaccgcacc tgtggcgccg gtgatgccgg
     ccacgatgcg tccggcgtag aggatc
//