LOCUS pNEB206a 2722 bp DNA circular 18-OCT-2007 DEFINITION Cloning vector pNEB206a, complete sequence. ACCESSION urn.local...d-cuvevjd KEYWORDS . SOURCE Cloning vector pNEB206a ORGANISM Cloning vector pNEB206a other sequences; artificial sequences; vectors. REFERENCE 1 (bases 1 to 2722) AUTHORS Bitinaite,J., Rubino,M., Varma,K.H., Schildkraut,I., Vaisvila,R. and Vaiskunaite,R. TITLE USER friendly DNA engineering and cloning method by uracil excision JOURNAL Nucleic Acids Res 35 (6), 1992-2002 (2007) PUBMED 17341463 REFERENCE 2 (bases 1 to 2722) AUTHORS Bitinaite,J. TITLE Direct Submission JOURNAL Submitted (18-OCT-2007) Research Department, New England Biolabs, 240 County Road, Ipswich, MA 01938, USA COMMENT The plasmid is supplied in a linearized form 2706 bp in length (with bp 438..453 excised from the circular form), including two flanking, noncomplementary 8-base 3' overhangs at the intended cloning site. This form consists of bases (454..2722,1..437) where 430..437 and 454..461 are single stranded. FEATURES Location/Qualifiers source 1..2722 /organism="Cloning vector pNEB206a" /mol_type="other DNA" gene complement(146..505) /gene="lacZalpha" CDS complement(146..505) /gene="lacZalpha" /codon_start=1 /product="beta-galactosidase alpha fragment" /translation="MTMITPSFLQGLNAEETSRGSSRLSLSLIKARLSSNSLAVVLQR RDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCG ISHRIWCTLSTICSDAA" misc_feature 396..488 /gene="lacZalpha" /note="multiple cloning site (EcoRI-HindIII); top strand nt 438-461 and bottom strand nt 430-453 deleted by XbaI + N.BbvCIB digestion" -10_signal complement(550..555) /note="Plac promoter (drives lacZalpha; counter-clockwise); TATGTT" -35_signal complement(574..579) /note="Plac promoter (drives lacZalpha; counter-clockwise); TTTACA" protein_bind complement(599..611) /note="CAP protein binding site" /bound_moiety="catabolite activator protein (CAP)" rep_origin complement(903..1491) /note="pMB1 origin of replication (counter-clockwise) (RNAII -35 to RNA/DNA switch point)" misc_RNA complement(903..1455) /note="RNAII transcript (complementary strand)" -35_signal 1309..1314 /note="RNAI promoter (clockwise); TTGAAG" -10_signal 1331..1336 /note="RNAI promoter (clockwise); GCTACA" misc_RNA 1345..1452 /note="RNAI transcript" -10_signal complement(1465..1470) /note="RNAII promoter (counter-clockwise); CGTAAT" -35_signal complement(1486..1491) /note="RNAII promoter (counter-clockwise); TTGAGA" gene complement(1662..2522) /gene="bla" CDS complement(1662..2522) /gene="bla" /note="ampR (confers resistance to ampicillin)" /codon_start=1 /product="beta-lactamase" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVE YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL DRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA EIGASLIKHW" sig_peptide complement(2454..2522) /gene="bla" /note="required for secretion to the periplasm; cleaved off to form the mature beta-lactamase protein." -10_signal complement(2566..2571) /note="bla promoter (counter-clockwise); GAGACA; transcript start 2557 (complementary strand)" -35_signal complement(2587..2592) /note="bla promoter (counter-clockwise); TTCAAA" ORIGIN 1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca 61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg 121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc 181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc 241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat 301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt 361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt cgagctcagg cgcgccttaa 421 ttaagctgag ggaaagtcta gaggatcctc tagatgtctc ctcagcgttt aaaccctgca 481 ggaagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca 541 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg 601 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg 661 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc 721 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta 781 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag 841 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg 901 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg 961 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg 1021 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga 1081 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc 1141 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt 1201 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact 1261 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg 1321 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt 1381 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt 1441 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct 1501 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg 1561 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt 1621 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt 1681 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc 1741 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg 1801 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc 1861 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg 1921 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca 1981 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga 2041 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct 2101 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg 2161 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca 2221 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata 2281 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct 2341 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact 2401 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa 2461 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc 2521 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga 2581 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga 2641 aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg 2701 cgtatcacga ggccctttcg tc //