LOCUS pBR322 4361 bp DNA circular 18-OCT-2007 DEFINITION Cloning vector pBR322, complete sequence. ACCESSION VERSION KEYWORDS . SOURCE Cloning vector pBR322 ORGANISM Cloning vector pBR322 other sequences; artificial sequences; vectors. REFERENCE 1 (bases 1 to 4361) AUTHORS New England Biolabs. TITLE Direct Submission JOURNAL Submitted (18-OCT-2007) Research Department, New England Biolabs, 240 County Road, Ipswich, MA 01938, USA COMMENT 1. pBR322 beta-lactamase (bla, conferring ampicillin resistance) is transcribed in vitro and in vivo both from its natural promoter (P3) and from promoter P1, a pSC101 promoter artificially juxtaposed with the bla sequence during construction of the pBR322 plasmid. (Ref: Brosius et al, J Biol Chem 257, 9205-9210 (1982)). 2. Corrects sequence error in tet gene deriving from the original pSC101 sequence, present in GenBank entry (see accession J01749) and NEB sequences dated 6/6/02 and earlier, based on sequencing of pBR322 and pACYC184 at NEB in 2007 (Mathieu-Sheltry, V. and Cantin, E., unpublished). Correction is C1134T, representing a Thr350Ile correction in the expressed protein. FEATURES Location/Qualifiers source 1..4361 /organism="Cloning vector pBR322" /mol_type="other DNA" -35_signal 10..15 /note="tet (P2) promoter (clockwise); TTGACA" -10_signal 33..38 /note="tet (P2) promoter (clockwise); TTTAAT; transcript start 45" -10_signal complement(44..49) /note="P1 promoter (counter-clockwise); TAAACT" -35_signal complement(64..75) /note="P1 promoter -35 sequence region; transcript starts 36, 37 (complementary strand)" gene 86..1276 /gene="tet" CDS 86..1276 /gene="tet" /note="tetR (confers resistance to tetracycline)" /codon_start=1 /product="tetracycline efflux protein, class C" /translation="MKSNNALIVILGTVTLDAVGIGLVMPVLPGLLRDIVHSDSIASH YGVLLALYALMQFLCAPVLGALSDRFGRRPVLLASLLGATIDYAIMATTPVLWILYAG RIVAGITGATGAVAGAYIADITDGEDRARHFGLMSACFGVGMVAGPVAGGLLGAISLH APFLAAAVLNGLNLLLGCFLMQESHKGERRPMPLRAFNPVSSFRWARGMTIVAALMTV FFIMQLVGQVPAALWVIFGEDRFRWSATMIGLSLAVFGILHALAQAFVTGPATKRFGE KQAIIAGMAADALGYVLLAFATRGWMAFPIMILLASGGIGMPALQAMLSRQVDDDHQG QLQGSLAALTSLTSIIGPLIVTAIYAASASTWNGLAWIVGAALYLVCLPALRRGAWSR ATST" RBS 1905..1909 /note="rop RBS" gene 1915..2106 /gene="rop" CDS 1915..2106 /gene="rop" /exception="alternative start codon" /codon_start=1 /transl_table=11 /product="ROP protein" /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDH ADELYRSCLARFGDDGENL" rep_origin complement(2534..3122) /note="pMB1 origin of replication (counter-clockwise) (RNAII -35 to RNA/DNA switch point)" misc_RNA complement(2534..3086) /note="RNAII transcript (complementary strand)" -35_signal 2940..2945 /note="RNAI promoter (clockwise); TTGAAG" -10_signal 2962..2967 /note="RNAI promoter (clockwise); GCTACA" misc_RNA 2976..3083 /note="RNAI transcript" -10_signal complement(3096..3101) /note="RNAII promoter (counter-clockwise); CGTAAT" -35_signal complement(3117..3122) /note="RNAII promoter (counter-clockwise); TTGAGA" gene complement(3293..4153) /gene="bla" CDS complement(3293..4153) /gene="bla" /note="ampR (confers resistance to ampicillin)" /codon_start=1 /product="beta-lactamase" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVE YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL DRWEPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA EIGASLIKHW" sig_peptide complement(4085..4153) /gene="bla" /note="required for secretion to the periplasm; cleaved off to form the mature beta-lactamase protein." RBS complement(4161..4165) /note="bla RBS" -10_signal complement(4197..4202) /note="bla (P3) promoter (counter-clockwise); GAGACA; transcript start 4188 (complementary strand)" -35_signal complement(4218..4223) /note="bla (P3) promoter (counter-clockwise); TTCAAA" BASE COUNT 983 a 1209 c 1134 g 1035 t ORIGIN 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 tcattggacc 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 //