LOCUS       SYNPBR322               4361 bp    DNA     circular SYN 30-SEP-2008
DEFINITION  Cloning vector pBR322, complete sequence.
ACCESSION   J01749 K00005 L08654 M10282 M10283 M10286 M10356 M10784 M10785
            M10786 M33694 V01119
VERSION     J01749.1
KEYWORDS    ampicillin resistance; beta-lactamase; cloning vector; drug
            resistance protein; origin of replication; plasmid; tetracycline
SOURCE      Cloning vector pBR322
  ORGANISM  Cloning vector pBR322
            other sequences; artificial sequences; vectors.
REFERENCE   1  (bases 1 to 3; 3259 to 4361)
  AUTHORS   Sutcliffe,J.G.
  TITLE     Nucleotide sequence of the ampicillin resistance gene of
            Escherichia coli plasmid pBR322
  JOURNAL   Proc. Natl. Acad. Sci. U.S.A. 75 (8), 3737-3741 (1978)
   PUBMED   358200
REFERENCE   2  (bases 1 to 4361)
  AUTHORS   Sutcliffe,J.G.
  TITLE     Complete nucleotide sequence of the Escherichia coli plasmid pBR322
  JOURNAL   Cold Spring Harb. Symp. Quant. Biol. 43 (Pt 1), 77-90 (1979)
   PUBMED   383387
REFERENCE   3  (bases 1500 to 2300)
  AUTHORS   Reed,R.R., Young,R.A., Steitz,J.A., Grindley,N.D. and Guyer,M.S.
  TITLE     Transposition of the Escherichia coli insertion element gamma
            generates a five-base-pair repeat
  JOURNAL   Proc. Natl. Acad. Sci. U.S.A. 76 (10), 4882-4886 (1979)
   PUBMED   388421
REFERENCE   4  (bases 2207 to 2265)
  AUTHORS   Covarrubias,L., Cervantes,L., Covarrubias,A., Soberon,X.,
            Vichido,I., Blanco,A., Kupersztoch-Portnoy,Y.M. and Bolivar,F.
  TITLE     Construction and characterization of new cloning vehicles. V.
            Mobilization and coding properties of pBR322 and several deletion
            derivatives including pBR327 and pBR328
  JOURNAL   Gene 13 (1), 25-35 (1981)
   PUBMED   6263753
REFERENCE   5  (bases 2000 to 2500)
  AUTHORS   Marians,K.J., Soeller,W. and Zipursky,S.L.
  TITLE     Maximal limits of the Escherichia coli replication factor Y
            effector site sequences in pBR322 DNA
  JOURNAL   J. Biol. Chem. 257 (10), 5656-5662 (1982)
   PUBMED   6279609
REFERENCE   6  (bases 1 to 80; 4151 to 4229; 4349 to 4361)
  AUTHORS   Brosius,J., Cate,R.L. and Perlmutter,A.P.
  TITLE     Precise location of two promoters for the beta-lactamase gene of
            pBR322. S1 mapping of ribonucleic acid isolated from Escherichia
            coli or synthesized in vitro
  JOURNAL   J. Biol. Chem. 257 (15), 9205-9210 (1982)
   PUBMED   6178738
REFERENCE   7  (bases 4241 to 4343)
  AUTHORS   Van Dyke,M.W., Hertzberg,R.P. and Dervan,P.B.
  TITLE     Map of distamycin, netropsin, and actinomycin binding sites on
            heterogeneous DNA: DNA cleavage-inhibition patterns with
  JOURNAL   Proc. Natl. Acad. Sci. U.S.A. 79 (18), 5470-5474 (1982)
   PUBMED   6291045
REFERENCE   8  (bases 584 to 709)
  AUTHORS   Peden,K.W. and Nathans,D.
  TITLE     Local mutagenesis within deletion loops of DNA heteroduplexes
  JOURNAL   Proc. Natl. Acad. Sci. U.S.A. 79 (23), 7214-7217 (1982)
   PUBMED   6760191
REFERENCE   9  (bases 373 to 649)
  AUTHORS   Peden,K.W.
  TITLE     Revised sequence of the tetracycline-resistance gene of pBR322
  JOURNAL   Gene 22 (2-3), 277-280 (1983)
   PUBMED   6307828
REFERENCE   10  (bases 132 to 181)
  AUTHORS   Watabe,H., Iino,T., Kaneko,T., Shibata,T. and Ando,T.
  TITLE     A new class of site-specific endodeoxyribonucleases. Endo.Sce I
            isolated from a eukaryote, Saccharomyces cerevisiae
  JOURNAL   J. Biol. Chem. 258 (8), 4663-4665 (1983)
   PUBMED   6300094
REFERENCE   11  (bases 368 to 581)
  AUTHORS   Livneh,Z.
  TITLE     Directed mutagenesis method for analysis of mutagen specificity:
            application to ultraviolet-induced mutagenesis
  JOURNAL   Proc. Natl. Acad. Sci. U.S.A. 80 (1), 237-241 (1983)
   PUBMED   6337373
REFERENCE   12  (bases 2627 to 2682; 2781 to 2828)
  AUTHORS   Mascharak,P.K., Sugiura,Y., Kuwahara,J., Suzuki,T. and Lippard,S.J.
  TITLE     Alteration and activation of sequence-specific cleavage of DNA by
            bleomycin in the presence of the antitumor drug
  JOURNAL   Proc. Natl. Acad. Sci. U.S.A. 80 (22), 6795-6798 (1983)
   PUBMED   6196777
REFERENCE   13  (bases 4276 to 4336)
  AUTHORS   Schultz,P.G. and Dervan,P.B.
  TITLE     Sequence-specific double-strand cleavage of DNA by
            penta-N-methylpyrrolecarboxamide-EDTA X Fe(II)
  JOURNAL   Proc. Natl. Acad. Sci. U.S.A. 80 (22), 6834-6837 (1983)
   PUBMED   6417654
REFERENCE   14  (bases 518 to 528)
  AUTHORS   Sutcliffe,J.G.
  JOURNAL   Unpublished
REFERENCE   15  (bases 2395 to 2495)
  AUTHORS   Fuller,R.S., Funnell,B.E. and Kornberg,A.
  TITLE     The dnaA protein complex with the E. coli chromosomal replication
            origin (oriC) and other DNA sites
  JOURNAL   Cell 38 (3), 889-900 (1984)
   PUBMED   6091903
REFERENCE   16  (bases 2729 to 2731)
  AUTHORS   Lathe,R., Kieny,M.P., Skory,S. and Lecocq,J.P.
  TITLE     Linker tailing: unphosphorylated linker oligonucleotides for
            joining DNA termini
  JOURNAL   DNA 3 (2), 173-182 (1984)
   PUBMED   6327214
REFERENCE   17  (bases 2729 to 2730)
  AUTHORS   Heusterspreute,M. and Davison,J.
  TITLE     Restriction site bank vectors. II. DNA sequence analysis of plasmid
  JOURNAL   DNA 3 (3), 259-268 (1984)
   PUBMED   6086259
REFERENCE   18  (bases 2113 to 2186; 2348 to 2415)
  AUTHORS   Abarzua,P., Soeller,W. and Marians,K.J.
  TITLE     Mutational analysis of primosome assembly sites. I. Distinct
            classes of mutants in the pBR322 Escherichia coli factor Y DNA
            effector sequences
  JOURNAL   J. Biol. Chem. 259 (22), 14286-14292 (1984)
   PUBMED   6209275
REFERENCE   19  (bases 2348 to 2415)
  AUTHORS   Soeller,W., Abarzua,P. and Marians,K.J.
  TITLE     Mutational analysis of primosome assembly sites. II. Role of
            secondary structure in the formation of active sites
  JOURNAL   J. Biol. Chem. 259 (22), 14293-14300 (1984)
   PUBMED   6150042
REFERENCE   20  (bases 1 to 4361)
  AUTHORS   Van Dyke,M.M. and Dervan,P.B.
  TITLE     Echinomycin binding sites on DNA
  JOURNAL   Science 225 (4667), 1122-1127 (1984)
   PUBMED   6089341
REFERENCE   21  (sites)
  AUTHORS   Pouwels,P.H., Enger-Valk,B.E. and Brammar,W.J.
  JOURNAL   (in) CLONING VECTORS. Elsevier Scientific Publishing, Amsterdam
  REMARK    Vector I-A-iv-1
REFERENCE   22  (bases 1 to 4361)
  AUTHORS   Watson,N.
  TITLE     A new revision of the sequence of plasmid pBR322
  JOURNAL   Gene 70 (2), 399-403 (1988)
   PUBMED   3063608
REFERENCE   23  (sites)
  AUTHORS   Gilbert,W.
  TITLE     Obtained from VecBase 3.0
  JOURNAL   Unpublished
COMMENT     On or before Apr 4, 2002 this sequence version replaced L08654.1,
            The circular sequence is numbered such that 0 is the middle of the
            unique EcoRI site and the count increases first through the tet
            genes, the pMB1 material, and finally through the Tn3 region.
            Plasmid pBR322 contains ampicillin and tetracycline resistance
            genes.  The ampicillin resistance gene (amp-r) is a penicillin
            beta-lactamase. Promoters P1 and P3 are for the beta-lactamase
            gene. P3 is the natural promoter, and P1 is artificially created by
            the ligation of two different DNA fragments to create pBR322. P2 is
            in the same region as P1, but it is on the opposite strand and
            initiates transcription in the direction of the tetracycline
            resistance gene.
            Mutational studies in the primosome assembly sites indicate four
            types of mutations:  Class I having no effect on the activities
            elicited by the DNA site and the bases involved are probably
            spacers; Class II requiring higher Mg-2+ concentrations than the
            wild-type to be fully activated as factor Y ATPase effectors; Class
            III co-inactivating both the ATPase effector and DNA replication
            template activity of the site, indicating that they probably
            represent essential contact points between factor Y and the DNA;
            Class IV having a replication template activity intermediate that
            of class III and class II mutant DNAs.
            Specific sites within or near the origins of replication are
            recognized by dnaA protein.  Without dnaA binding to the origin of
            replication chromosomal replication is not possible (15). pBR322
            DNA contains two separate regions on opposite strands and close to
            the origin of replication which, when in single-stranded form, can
            act as effectors for the ATPase activity of E.coli replication
            factor Y (5).  Small fragments of DNA containing these sites when
            cloned in an f1 phage vector act as origins of DNA replication
            allowing the formation of complementary double-stranded DNA in
            rifampicin-resistant, dna(B,G,C)-dependent fashion in vitro (5).
            The biological activity of echinomycin is thought to be related to
            the formation of complexes by intercalating with cellular DNA (20).
            Complete source information:
            Plasmid pBR322 from E.coli (2),(1),(3),(6),(11),(8),(5),(7),(12),
            (13),(10),(9),(14),(18),(19),(15),(20),(16); pBR322 DNA in pXf3
            The following data and their annotation were supplied by Will
            Gilbert under the auspices of the Curator Program.
                             GenBank(50):pSC101C, GenBank(50):Trn3
                             VecBase(3):pBR325, VecBase(3):pBR327,
                             VecBase(3):pAT153, VecBase(3):pUC7,
                             VecBase(3):PiVX, VecBase(3):PiAN7,
                             VecBase(3):pSP65, VecBase(3):pGEM1,
                             VecBase(3):pGEM3, VecBase(3):pGEM4,
                             VecBase(3):pLBU3, VecBase(3):pTrS3,
                             VecBase(3):pSV2Cat, VecBase(3):M13mp9,
                             VecBase(3):pV34, VecBase(3):pKTH601,
                             VecBase(3):pKTH605, VecBase(3):pKTH606,
                             VecBase(3):YIp5, VecBase(3):YRp17,
                             VecBase(3):pSP6T3, VecBase(3):pSP6T719,
                             VecBase(3):pT713, VecBase(3):pT7T318,
                             VecBase(3):pT7T3A18, VecBase(3):pT7T3A19,
                             VecBase(3):pEX2, VecBase(3):pEX3,
                             VecBase(3):pACYC177, VecBase(3):pKO1,
                             VecBase(3):pKM2, VecBase(3):pMBL1,
                             VecBase(3):pMC1511, VecBase(3):pMC1871,
                             VecBase(3):pUR278, VecBase(3):pUR288,
                             VecBase(3):pUR290, VecBase(3):pUR291,
FEATURES             Location/Qualifiers
     source          1..4361
                     /organism="Cloning vector pBR322"
                     /mol_type="other DNA"
                     /tissue_lib="ATCC 31344, ATCC 37017"
     source          1..1762
                     /organism="Plasmid pSC101"
                     /mol_type="other DNA"
     misc_binding    24..27
     regulatory      complement(27..33)
                     /note="promoter P1 (6)"
     misc_binding    39..42
     regulatory      43..49
                     /note="promoter P2 (6)"
     misc_binding    53..56
     misc_binding    67..70
     misc_binding    80..83
     gene            86..1276
     CDS             86..1276
                     /product="tetracycline resistance protein"
     misc_feature    complement(141..142)
                     /note="Endo.Sce I cleavage site coordinated with site at
                     base 146 (10)"
     misc_feature    146..147
                     /note="Endo.Sce I cleavage site coordinated with site at
                     base 142 (10)"
     misc_binding    411..414
     misc_difference 426
     misc_binding    469..472
     old_sequence    526..528
     repeat_region   complement(1515..1519)
                     /note="gamma-delta insertion target sequence"
     misc_feature    1636..1762
                     /note="from pSC101 (bp 1860-1986)"
     repeat_region   complement(1788..1792)
                     /note="gamma-delta insertion target sequence"
     misc_difference 1891..1892
     old_sequence    1892..1893
     regulatory      1905..1910
     regulatory      1905..1909
                     /note="Shine-Dalgarno sequence"
     misc_difference 1913..1914
     old_sequence    1914..1915
     CDS             1915..2106
                     /product="ROP protein"
     misc_feature    2011..2167
                     /note="H-strand Y effector site"
     repeat_region   complement(2245..2249)
                     /note="gamma-delta insertion target sequence"
     misc_feature    complement(2351..2414)
                     /note="L-strand Y effector site"
     misc_binding    2439..2447
     rep_origin      2535
     old_sequence    2729..2730
                     /note="revision according to (17)"
     old_sequence    2729
     old_sequence    2730
                     /note="revision according to (16)"
     mobile_element  3148..4361
     repeat_region   3148..3185
                     /note="corresponds to one of the 38bp repeats found in Tn3
                     (bp 1-38 and complement (4920-4957))"
     gene            complement(3293..4153)
     CDS             complement(3293..4153)
     sig_peptide     complement(4085..4153)
     mat_peptide     complement(3296..4084)
     regulatory      complement(4161..4165)
                     /note="Shine-Dalgarno sequence"
     regulatory      complement(4188..4194)
                     /note="promoter P3 (6)"
     misc_binding    complement(4268..4271)
     misc_binding    complement(4280..4283)
     misc_binding    complement(4285..4288)
     misc_binding    complement(4296..4299)
     misc_binding    complement(4311..4314)
     misc_binding    complement(4317..4320)
     misc_binding    complement(4331..4334)
BASE COUNT          983 a         1210 c         1134 g         1034 t
        1 ttctcatgtt tgacagctta tcatcgataa gctttaatgc ggtagtttat cacagttaaa
       61 ttgctaacgc agtcaggcac cgtgtatgaa atctaacaat gcgctcatcg tcatcctcgg
      121 caccgtcacc ctggatgctg taggcatagg cttggttatg ccggtactgc cgggcctctt
      181 gcgggatatc gtccattccg acagcatcgc cagtcactat ggcgtgctgc tagcgctata
      241 tgcgttgatg caatttctat gcgcacccgt tctcggagca ctgtccgacc gctttggccg
      301 ccgcccagtc ctgctcgctt cgctacttgg agccactatc gactacgcga tcatggcgac
      361 cacacccgtc ctgtggatcc tctacgccgg acgcatcgtg gccggcatca ccggcgccac
      421 aggtgcggtt gctggcgcct atatcgccga catcaccgat ggggaagatc gggctcgcca
      481 cttcgggctc atgagcgctt gtttcggcgt gggtatggtg gcaggccccg tggccggggg
      541 actgttgggc gccatctcct tgcatgcacc attccttgcg gcggcggtgc tcaacggcct
      601 caacctacta ctgggctgct tcctaatgca ggagtcgcat aagggagagc gtcgaccgat
      661 gcccttgaga gccttcaacc cagtcagctc cttccggtgg gcgcggggca tgactatcgt
      721 cgccgcactt atgactgtct tctttatcat gcaactcgta ggacaggtgc cggcagcgct
      781 ctgggtcatt ttcggcgagg accgctttcg ctggagcgcg acgatgatcg gcctgtcgct
      841 tgcggtattc ggaatcttgc acgccctcgc tcaagccttc gtcactggtc ccgccaccaa
      901 acgtttcggc gagaagcagg ccattatcgc cggcatggcg gccgacgcgc tgggctacgt
      961 cttgctggcg ttcgcgacgc gaggctggat ggccttcccc attatgattc ttctcgcttc
     1021 cggcggcatc gggatgcccg cgttgcaggc catgctgtcc aggcaggtag atgacgacca
     1081 tcagggacag cttcaaggat cgctcgcggc tcttaccagc ctaacttcga tcactggacc
     1141 gctgatcgtc acggcgattt atgccgcctc ggcgagcaca tggaacgggt tggcatggat
     1201 tgtaggcgcc gccctatacc ttgtctgcct ccccgcgttg cgtcgcggtg catggagccg
     1261 ggccacctcg acctgaatgg aagccggcgg cacctcgcta acggattcac cactccaaga
     1321 attggagcca atcaattctt gcggagaact gtgaatgcgc aaaccaaccc ttggcagaac
     1381 atatccatcg cgtccgccat ctccagcagc cgcacgcggc gcatctcggg cagcgttggg
     1441 tcctggccac gggtgcgcat gatcgtgctc ctgtcgttga ggacccggct aggctggcgg
     1501 ggttgcctta ctggttagca gaatgaatca ccgatacgcg agcgaacgtg aagcgactgc
     1561 tgctgcaaaa cgtctgcgac ctgagcaaca acatgaatgg tcttcggttt ccgtgtttcg
     1621 taaagtctgg aaacgcggaa gtcagcgccc tgcaccatta tgttccggat ctgcatcgca
     1681 ggatgctgct ggctaccctg tggaacacct acatctgtat taacgaagcg ctggcattga
     1741 ccctgagtga tttttctctg gtcccgccgc atccataccg ccagttgttt accctcacaa
     1801 cgttccagta accgggcatg ttcatcatca gtaacccgta tcgtgagcat cctctctcgt
     1861 ttcatcggta tcattacccc catgaacaga aatccccctt acacggaggc atcagtgacc
     1921 aaacaggaaa aaaccgccct taacatggcc cgctttatca gaagccagac attaacgctt
     1981 ctggagaaac tcaacgagct ggacgcggat gaacaggcag acatctgtga atcgcttcac
     2041 gaccacgctg atgagcttta ccgcagctgc ctcgcgcgtt tcggtgatga cggtgaaaac
     2101 ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc
     2161 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggcgc agccatgacc
     2221 cagtcacgta gcgatagcgg agtgtatact ggcttaacta tgcggcatca gagcagattg
     2281 tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc
     2341 gcatcaggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     2401 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     2461 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     2521 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     2581 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     2641 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     2701 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     2761 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     2821 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     2881 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     2941 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc
     3001 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     3061 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     3121 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     3181 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     3241 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     3301 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     3361 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     3421 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     3481 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     3541 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     3601 ccattgctgc aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     3661 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     3721 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     3781 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     3841 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     3901 cggcgtcaac acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     3961 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     4021 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     4081 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     4141 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     4201 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     4261 catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct
     4321 ataaaaatag gcgtatcacg aggccctttc gtcttcaaga a