Fig. 3.
Verification of rrp1 deletion by allelic exchange replacement and analysis of growth rate of Bb5A13-wt and Bb5A13-Δrrp1. To confirm allelic exchange replacement was successful, PCR (A), Southern hybridization (B) and immunoblot analyses (C) were performed. (A) shows a schematic of the chromosome after allelic exchange, in which rrp1 (BB0419) has been replaced by the spectinomycin/streptomycin resistance cassette (SpecR). Primers are indicated by numbers and listed in Table S2. Numbers above each lane in the ethidium bromide-stained gel indicate primer combinations. In (B), EcoRI-restricted genomic DNA was fractionated, blotted and screened with a radiolabeled rrp1 probe. In (C), cell lysates were screened with Rrp1 antiserum. In (D), Bb5A13-wt and Bb5A13-Δrrp1 cells were cultivated at 25°C, 33°C and 37°C in BSK-H complete media. All methods were as described in the text.