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Construction and evaluation of a Clostridium thermocellum ATCC 27405 whole-genome oligonucleotide microarray

Applied Biochemistry and Biotechnology volume 137, pages 663–674 (2007)Cite this article

Abstract

Clostridium thermocellum is an anaerobic, thermophilic bacterium that can directly convert cellulosic substrates into ethanol. Microarray technology is a powerful tool to gain insights into cellular processes by examining gene expression under various physiological states. Oligonucleotide microarray probes were designed for 96.7% of the 3163 C. thermocellum ATCC 27405 candidate protein-encoding genes and then a partial-genome microarray containing 70 C. thermocellum specific probes was constructed and evaluated. We detected a signal-to-noise ratio of three with as little as 1.0 ng of genomic DNA and only low signals from negative control probes (nonclostridial DNA), indicating the probes were sensitive and specific. In order to further test the specificity of the array we amplified and hybridized 10 C. thermocellum polymerase chain reaction products that represented different genes and found gene specific hybridization in each case. We also constructed a whole-genome microarray and prepared total cellular RNA from the same point in early-logarithmic growth phase from two technical replicates during cellobiose fermentation. The reliability of the microarray data was assessed by cohybridization of labeled complementary DNA from the cellobiose fermentation samples and the pattern of hybridization revealed a linear correlation. These results taken together suggest that our oligonucleotide probe set can be used for sensitive and specific C. thermocellum transcriptomic studies in the future.

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Author notes

  1. Zhili He

    Present address: Institute of Environmental Genomics, Department of Botany and Microbiology, University of Oklahoma, Norman, OK

Affiliations

  1. Microbial Ecology and Physiology Group, Oak Ridge National Laboratory, Oak Ridge, TN

    Steven D. Brown & Zhili He

  2. Bioconversion Science and Technology Group, Biosciences Division, Oak Ridge National Laboratory, Oak Ridge, TN

    Babu Raman, Catherine K. McKeown & Jonathan R. Mielenz

  3. Department of Biology, Xavier University of Louisiana, New Orleans, LA

    Shubha P. Kale

Authors
  1. Steven D. Brown
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  2. Babu Raman
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  3. Catherine K. McKeown
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  4. Shubha P. Kale
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  5. Zhili He
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  6. Jonathan R. Mielenz
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Correspondence to Jonathan R. Mielenz.

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Brown, S.D., Raman, B., McKeown, C.K. et al. Construction and evaluation of a Clostridium thermocellum ATCC 27405 whole-genome oligonucleotide microarray. Appl Biochem Biotechnol 137, 663–674 (2007). https://doi.org/10.1007/s12010-007-9087-6

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  • DOI: https://doi.org/10.1007/s12010-007-9087-6

Index Entries

  • Biomass
  • cellulose
  • ethanol
  • fermentation
  • transcriptomics