K-122. Gene Expression Profiles of Clostridium botulinum Using Microarray

Y-H. Lin;
California State Polytechnic Univ., Pomona, CA.

Background: Clostridium botulinum is a gram-positive anaerobic bacterium known to produce botulinum neurotoxin. The expression of neurotoxin is temporally regulated by the transcription regulator, BotR. However, it is still unclear how toxin expression correlates with cellular growth and physiology. Microarray is widely used for the generation of global gene expression profiles. From the expression profile we would be able to determine how toxin formation is correlated to cellular events. Our goal is to study global gene expression profiles of C. botulinum Type A using DNA microarray. Methods: RNA samples were purified from cells harvested at 7 time points post-inoculation. RNA samples were reverse-transcribed into cDNA with aa-dUTP. Then cDNA samples were labeled with Cy3 or Cy5 fluorescent dyes and mixed with the hybridization mixture, then applied onto the microarray slide. The slides were scanned by a fluorescent scanner to generate fluorescent images. From these images, raw gene expression data were generated, and those data were further analyzed using the TIGR TM4 software. Results: Gene expression profiles of C. botulinum cultures harvested throughout the growth phases were analyzed by microarray using pooled RNAs as a control. The expression patterns over the growth period were categorized into several basic gene expression patterns and were summarized into two major trends of expression. The first major trend shows genes start with higher expression levels then decrease after the exponential growth phase. Genes important for metabolism and cell division exemplify this expression trend. The second trend shows genes with low expression levels in the early stages of the growth increase over time. This trend includes genes from sporulation and botulinum neurotoxin complex. Conclusion: We report the global gene expression profiles of C. botulinum during the first 11 hours of growth until the stationary growth phase. Several temporal gene expression patterns were identified and analyzed. This data will provide a basis for further growth-phases study to confirm temporally regulated genes which may be essential in growth, sporulation, and neurotoxin formation in C. botulinum.