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ID   PATH20     preliminary; circular DNA; SYN; 3793 BP.
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AC   ATCC37700;
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DT   01-JUL-1993 (Rel. 7, Created)
DT   01-JUL-1995 (Rel. 12, Last updated, Version 1)
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DE   E. coli plasmid vector pATH20 - 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]
RC   pATH1 from pKRS101 & linker
RC   pATH2 from pATH1 & linker
RC   pATH3 from pATH1 & linker
RC   pATH10 from pATH1 & pUC8
RC   pATH11 from pATH1 & pUC8
RC   pATH20 from pATH1 & pUC18
RC   pATH21 from pATH1 & pUC18
RC   pATH22 from pATH1 & pUC18
RC   pATH23 from pATH1 & pUC18
RA   Koerner T.J., Hill J.E., Myers A.M., Tzagoloff A.;
RT   "High-expression vectors with multiple cloning sites for construction
RT   of trpE-fusion genes: pATH vectors";
RL   Meth. Enzymol. 194:477-490(1991).
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CC   Created by Moore, July 1995, under contract with NCBI.
CC   The nucleotides remaining in the codon after cutting the vector with
CC   the given enzyme are:BamHI-1, ClaI-?, EcoRI-1, HindIII-1, KpnI-2,
CC   PstI-2, SacI-2, SalI-1, SmaI-1, SphI-2, XbaI-1, XmaI-2.
CC   Restriction digests of the clone give the following sizes (kb):
CC   BamHI--3.85; EcoRI--3.85; HindIII--3.85. (ATCC staff)
CC   Nucleotide 1 is the first in the trp fragment, at the beginning of a
CC   PvuII site 261 nucleotides upstream of the transcription start site.
CC   The multiple cloning site follows codon 323 of trpE.
CC   To prevent recircularization of the vector, the depositors recommend
CC   including 30 - 50 U of calf alkaline phosphatase in the restriction
CC   enzyme digestion used to prepare the vector.
CC   E. coli containing pATH vectors should be grown on medium
CC   supplemented with tryptophan to prevent expression. Storage as DNA at
CC   -20C is preferred; long term storage as cells may result in selection
CC   of non-expressing promoter mutations.
CC   Fusion proteins are produced in E.coli usually in an insoluble
CC   form which facilitates purification.Hybrid proteins larger than 90 kDa
CC   are not expressed as efficiently as shorter ones.
CC   This is one of a series of vectors (ATCC 37695-37703) designed to
CC   facilitate expression of an open reading frame as a trpE fusion
CC   protein.
CC   Medium is -1 1065 plus ampicillin (50 ug/ml) & tryptophan (10 ug/ml).
CC   NM (pATH20)
CC   CM (yes)
CC   NA (ds-DNA)
CC   TP (circular)
CC   ST ()
CC   TY (plasmid)
CC   SP (ATCC)
CC   HO (E.coli RR1)(E.coli)
CC   CP ()
CC   FN (cloning)
CC   SE ()
CC   PA (pATH1)(pUC18)(pUC8)
CC   BR ()
CC   OF ()
CC   OR ()
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FH   Key             Location/Qualifiers
FH
FT   misc_feature    0..0
FT                   /note="1. pATH1 remove AvaII-AvaII 222bp 2120..2342,
FT                   \ amp gene 3557bp
FT                   2. pUC8 AvaII-AvaII 222bp 1288..1510, amp gene
FT                   -> plasmid 3779bp [no PstI in amp]
FT                   1. plasmid remove SmaI-ClaI 37bp 1421..1458,
FT                   \ MCS 3742bp
FT                   blunt end:blunt end
FT                   2. pUC18 HindIII-EcoRI 51bp 400..451, MCS
FT                   blunt end:blunt end
FT                   -> pATH20 3793bp [no PstI in amp]"
FT   -               1..1420
FT                   /note="pATH1 1..1420 1420bp
FT                   SmaI =  CCC^GGG
FT                   HindIII = A^AGCTT"
FT   -               1421..1471
FT                   /note="pUC18 400..450 51bp
FT                   EcoRI = G^AATTC
FT                   ClaI = AT^CGAT"
FT   -               1472..2133
FT                   /note="pATH1 1458..2119 662bp
FT                   AvaII = G^GWCC"
FT   -               2134..2355
FT                   /note="pUC8 1288..1509 222bp
FT                   AvaII = G^GWCC"
FT   -               2356..3793
FT                   /note="pATH1 2342..3779 1438bp"
FT   misc_binding    0..0
FT                   /note="MCS BamHI-SmaI-SstI-EcoRI"
FT   misc_binding    0..0
FT                   /note="SIT unique BamHI-EcoRI-HindIII-KpnI-PstI-
FT                   SacI-SalI-SmaI-SphI-XbaI-XmaI"
FT   rep_origin      0..0
FT                   /note="ORI E. coli pMB1 (ColE1 and pBR322)"
FT   promoter        0..0
FT                   /note="PRO E. coli trp gene"
FT   CDS             0..0
FT                   /note="ANT E. coli beta-lactamase gene (bla)
FT                   ampicillin resistance gene (apr/amp)"
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SQ   Sequence 3793 BP; 929 A; 952 C; 958 G; 954 T; 0 other;
     cagctgtggt gtcatggtcg gtgatcgcta gggtgccgag cgcatctcga ctgcacggtg
     caccaatgct tctggcgtca ggtagttatt ggaaagctgt ggtatggctg tgcaggtcgt
     aaatcactgc ataactcgct gctgcctaag gcgcactccc gttctggata atgttttttg
     cgccgacatc ataacggttc tggcaaatat tctgaaatga gctgttgaca attaatcatc
     gaactagtta actagtacgc aagttcacgt aaaaagggta tcgacaatga aagcaatttt
     cgtactgaaa ggttggtggc gcacttcctg aaacgggcag tgtattcacc atgcgtaaag
     caatcagata cccagcccgc ctaatgagcg ggcttttttt tgaacaaaat tagagaataa
     caatgcaaac acaaaaaccg actctcgaac tgctaacctg cgaaggcgct tatcgcgaca
     atcccaccgc gctttttcac cagttgtgtg gggatcgtcc ggcaacgctg ctgctggaat
     ccgcagatat cgacagcaaa gatgatttaa aaagcctgct gctggtagac agtgcgctgc
     gcattacagc tttaggtgac actgtcacaa tccaggcact ttccggcaac ggcgaagccc
     tcctggcact actggataac gccctgcctg cgggtgtgga aagtgaacaa tcaccaaact
     gccgtgtgct gcgcttcccc cctgtcagtc cactgctgga tgaagacgcc cgcttatgct
     ccctttcggt ttttgacgct ttccgtttat tgcagaatct gttgaatgta ccgaaggaag
     aacgagaagc catgttcttc agcggcctgt tctcttatga ccttgtggcg ggatttgaag
     atttaccgca actgtcagcg gaaaataact gccctgattt ctgtttttat ctcgctgaaa
     cgctgatggt gattgaccat cagaaaaaaa gcacccgtat tcaggccagc ctgtttgctc
     cgaatgaaga agaaaaacaa cgtctcactg ctcgcctgaa cgaactacgt cagcaactga
     ccgaagccgc gccgccgctg ccagtggttt ccgtgccgca tatgcgttgt gaatgtaatc
     agagcgatga agagttcggt ggcgtagtgc gtttgttgca aaaagcgatt cgcgctggag
     aaattttcca ggtggtgcca tctcgccgtt tctctctgcc ctgcccgtca ccgctggcgg
     cctattacgt gctgaaaaag agtaatccca gcccgtacat gttttttatg caggataatg
     atttcaccct atttggcgcg tcgccggaaa gctcgctcaa gtatgatgcc accagccgcc
     agattgagat ccccgggcga gctcgaattc gagctcgccc agcttgcatg cctgcaggtc
     gactctagag gatccccggg taccgagctc gcgatgataa gctgtcaaac atgagaatta
     attcttgaag acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa
     taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     aagttctgct atgtggcgcg gtattatccc gtgttgacgc cgggcaagag caactcggtc
     gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     tacagcgtga gcattgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt ctccttacgc
     atctgtgcgg tatttcacac cgcatatggt gcactctcag tacaatctgc tctgatgccg
     catagttaag ccagtataca ctccgctatc gctacgtgac tgggtcatgg ctgcgccccg
     acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
     cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
     gaaacgcgcg agg
//