Chapter 17 : Genetic Manipulation of

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The genus is classified in the family of the phylum of . The genetic manipulation of clostridia is still in an early stage of development, but significant advances have been made in recent years, and this chapter emphasizes newly developed genetic methods and strategies. Certain physiological properties of clostridia are integral for genetic manipulation and gene expression. Clostridia are classified as proteolytic or saccharolytic depending on their preference for energy-yielding substrates. Conjugal transfer between the same and different species of certainly occurs in the environment and in the gastrointestinal tracts of mammals, but it is not used for routine genetic manipulation. This chapter describes the replicons, selective markers, and reporter genes most frequently used for construction of clostridial shuttle vectors. It also describes the latest developments in the field. The most comprehensive and detailed transcriptional analysis including the life cycle of has been performed in the E. T. Papoutsakis laboratory. This study described major cellular regulatory systems; sigma and sporulation factors, including activity assays for major sporulation factors of the canonical sets of genes from their regulons; assessment of expression intensities; and identification of putative histidine kinases that may phosphorylate Spo0A. The availability of genomic sequences provides insights into population genetics and evolution of . Ideally, the development of further genetic tools will foster fruitful collaborations within the clostridial community to understand this complex and difficult organism.

Citation: Bradshaw M, Johnson E. 2010. Genetic Manipulation of , p 238-261. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch17
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Image of FIGURE 1

Characteristic spindle morphology of and presence of endospores. The photograph shows a phase-contrast visual micrograph (750×) of a culture of type A.

Citation: Bradshaw M, Johnson E. 2010. Genetic Manipulation of , p 238-261. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch17
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Image of FIGURE 2

(A) Schematic presentation of pMTL80000-series modular plasmids, showing most available modules. (B) The MCS module. TT1, transcriptional terminator from downstream of the CD0164 ORF of 630; TT2, transcriptional terminator from the gene of . (Reprinted from reference with permission of the publisher.)

Citation: Bradshaw M, Johnson E. 2010. Genetic Manipulation of , p 238-261. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch17
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Generic image for table

Replicons used for construction of clostridial shuttle vectors

Citation: Bradshaw M, Johnson E. 2010. Genetic Manipulation of , p 238-261. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch17
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Antibiotic resistance genes for construction of clostridial shuttle vectors

Citation: Bradshaw M, Johnson E. 2010. Genetic Manipulation of , p 238-261. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch17
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Reporter genes used for construction of clostridial shuttle vectors

Citation: Bradshaw M, Johnson E. 2010. Genetic Manipulation of , p 238-261. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch17
Generic image for table

Numbering scheme for pMTL80000-series modular plasmids

Citation: Bradshaw M, Johnson E. 2010. Genetic Manipulation of , p 238-261. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch17
Generic image for table

Clostridial genome sequences

Citation: Bradshaw M, Johnson E. 2010. Genetic Manipulation of , p 238-261. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch17

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