Antibiotics were added to growth media at the following concentrations: ampicillin, 100 μg/ml; chloramphenicol, 10 μg/ml; erythromycin, 5 μg/ml; penicillin G, 0.03 or 0.09 μg/ml; and spectinomycin (SPC), 60 μg/ml. The isolation of chromosomal and plasmid

DNA, restriction enzyme analysis, and PCR were performed according to standard protocols [29]. PCR and RT-PCR primers used in this study are listed in Table 4. Table 4 Primers used in this study Primer Sequence [5′ → 3′] 16S RNA-E a TTAGCTAGTTGGTAGGGT 16S RNA-B a AATCCGGACAACGCTTGC 0943F ab CATTGGTATATGAGAGGCCAC 0943R ab CATTGTCGCCTTCTTTGTCAG 0944F ab ATGGTTTCATGATGAGTTTGATGT 0944R2 b ATTTTCCAGTCGTGGTCTTTG 0944R ac AL3818 TCCGTTTTTGGTTCATAGTCG 0945F ab CCGCACGCAGACCATATTG 0945R2 b ATTGGCACCGCTATCTACC 0945R ac CTGGTTGGATGTGGACGATC 1065F a GCTTGAAGCACGCATGACC Temozolomide mw 1065R a GCCGTCATGCACAGGATAC 1211F a CAGGTTTGTTAGCTGGGATG 1211R a ACGCCAAGTAGACGTTCGA 1622F a TAGCGTCAACCGTCCTGCT 1622R a ATCTCCCATACCGCCAGTG 1660F a TACCGCGTACGCAGATCG 1660R a GAATCAACACGTAGTCCGC

1941F a CCGGCTGATTATGACATGAG 1941R a TGCTTTCTCGGCAGCAGC 2501F a GTGGTGACAGCTGAAGATG 2501R a GTGGTGACAGCTGAAGATG 2820F a GCCTTGTCGCTTCGTGTG 2820R a ACTAAGACAACGGGCAGTC llo-1 d CGGGTACCAGGTAGAGCGGACATCCATTG llo-2 e GTTTTAGGATCCCCCGGGGGGTTTCACTCTCCTTCTAC llo-3 CCCGGGGGATCCTAAAACCGCTTAACACACACG llo-4 f GCGTCTAGATTCTTCCCCGACAGAATCTGC phoP-1 g CAGGATCCAGTTTTGGGTGCTCGTGC phoP-2 h TCGAATTCCTATCTACCATCTTCAGCTGTCAC phoP-3 h TCGAATTCGGACTTGAACTTGGAGCAG phoP-4 i CGTCCATGGTTACGTTCTCCATTTTATAACCG axyR-1 g CAGGATCCGGTAGCGATTAATTTTCACGAC selleck kinase inhibitor axyR-2 h TCGAATTCCTAATCATTGACTTCTTTCCTAGCAGA axyR-3 h TCGAATTCCTTATGCTAGTGAACTGGAATAC axyR-4

i CTCCCATGGCCGTAATCGTCTCATCGCTC Hly-1 g GCGGGATCCTGTAGAAGGAGAGTGAAACCCATG Hly-2 j GCGGTCGACACAATTATTCGATTGGATTATCTAC seq-1 CAGGAAACAGCTATGACCATG seq-2 ACTAATATAAGTGTAATAAAAACTAGCAT a Primers used for analysis of gene expression under stress conditions. b Primers used for PCR in cotranscription analysis. c Primer used for reverse transcription in cotranscription analysis. d The sequence in boldface type is the KpnI restriction enzyme site. e The underlined sequence is an overhang complementary to primer llo-3. f The sequence Cediranib (AZD2171) in boldface type is the XbaI restriction enzyme site. g The sequence in boldface type is the BamHI restriction enzyme site. h The sequence in boldface type is the EcoRI restriction enzyme site. i The sequence in boldface type is the NcoI restriction enzyme site. j The sequence in boldface type is the SalI restriction enzyme site. Construction and analysis of L. monocytogenes genomic libraries Two ~400-bp DNA fragments flanking the L. monocytogenes hly gene were amplified by PCR using strain EGD chromosomal DNA as the template. The primers used to amplify the hly 5′ flanking fragment were llo-1 and llo-2, and those for the 3′ fragment were llo-3 and llo-4.