mRNA levels of argB-20 and argB-37 in L. hongkongensis HLHK9 cells grown in 20uC and 37uC were compared. Total RNA was extracted from culture of L. hongkongensis HLHK9 (OD 600 of 0.6) grown in conditions described in proteomic analysis by using RNeasy kit (Qiagen) in combination with RNAprotect Bacteria Reagent (Qiagen) as described by the manufacturer. Genomic DNA was removed by DNase digestion using RNase-free DNase I (Roche). The total nucleic acid concentration and purity were estimated using A 260 /A 280 values measured by NanoDrop ND-1000 spectrophotometer (NanoDrop Technologies). Bacteria were harvested from three independent replicate cultures. cDNA was synthesized by RT using random hexamers and SuperScript III kit (Invitrogen) as described previously [53, 54] (Table S4) . Reactions were first incubated at 50uC for 2 min, followed by 95uC for 10 min in duplicate wells. Reactions were then thermal-cycled in 40 cycles of 95uC for 15 s and 60uC for 1 min. Absolute standard curve method was used for determination of transcript level for each gene. Standard curves were made by using serial dilutions from plasmids containing the target sequences with known quantities. Housekeeping gene RNA polymerase beta subunit, rpoB, was used as an internal control. Triplicate assays using RNAs extracted in three independent experiments confirmed that transcript levels of rpoB were not significantly different (P.0.05) at 20uC compared with 37uC (data not shown). The transcript levels of argB-20 and argB-37 were then normalized to that of rpoB. Triplicate assays using RNAs extracted in three independent experiments were performed for each target gene.