© 1995 • 540 pp., illus., appendices, index (volume 2 of a three-book set)
Plastic comb binding • ISBN  978-087969439-5

Foreword by Carl R. Woese, University of Illinois

The use of ribosomal RNA sequences as molecular chronometers hasled to the recognition of a group of prokaryotes, the Archaea, thatare phylogenetically distinct from both bacteria and eukaryotes.Although the Archaea are prokaryotes, lacking a nuclear membrane andpossessing a single circular chromosome, they possess severalmolecular properties with similarity to the eukaryotes such astranscription signals, transcription factors, chaperones, andhistones. Their unique phylogeny is underscored by a predilectionfor extreme environments which include high temperature, high salt,and strictly anaerobic conditions and the adaptive strategies thatthey have evolved to survive in these environments. These uniquebiomolecular adaptations have recently generated interest in theareas of genetic variability and genome evolution, extremelythermostable enzymes, such as DNA polymerases for PCR, methanogenesisas an alternative fuel source, anaerobic bioremediation, and use ofbacteriorhodopsin for production of a biocomputer chip or a model fortransmembrane ion translocation.

The Archaea occupy a pivotal phylogenetic position and offerextraordinary potential for biotechnology. One of the difficultiesfacing researchers, especially newcomers who wish to study theArchaea, has been the lack of a comprehensive source for thespecialized techniques required. As the field has evolved, solutionsto many problems have been discovered empirically and the specialtechniques have largely been disseminated either verbally or by meansof journal articles. The protocols in these three books are selectedto provide a detailed guide to experiments with themethanogenic, extremelyhalophilic, andthermophilic sulfur-utilizing Archaea,with overviews to highlight areas of future development. Theindividual protocols consist of an introduction describing thespecific applications of the techniques, step-by-step procedures forapplying the protocols, followed by any additional comments that willfacilitate successful application of the protocol. A feature ofresearch in this area is the interplay between microbiology,bioengineering, bio-chemistry, and molecular biology, and authorsfrom all of these fields have been selected to provide these threeconcise and comprehensive resources for scientists interested inconducting research on the Archaea.


• Volume 1: Halophilic Archaea

• Volume 2: MethanogenicArchaea

• Volume 3: Thermophilic Archaea


Methanogenic Archaea: An Overview (K.R. Sowers)

Growth and Identification: Protocol 1. Techniques for Anaerobic Growth (K.R. Sowers, K.M. Noll); Protocol 2. Growth of Methanogens on Solidified Medium (D.L. Tumbula et al.); Protocol 3. Plating Techniques for Extremely Thermophilic Methanogens (A. Rodabough et al.); Protocol 4. Growth of Methanosarcina spp. as Single Cells (K.R. Sowers); Protocol 5. Large-scale Culture Techniques (L. Daniels); Protocol 6. Large-scale Growth of Acetate- and Formate-utilizing Methanogens in a pH Auxostat (K.R. Sowers); Protocol 7. Short- and Long-term Maintenance of Stock Cultures (D.R. Boone); Protocol 8. Long-term Maintenance of Stock Cultures in Glycerol (D.L. Tumbula et al.); Protocol 9. Techniques for Monitoring Cell Growth (K.R. Sowers); Protocol 10. Immunologic Identification and Analysis (E. Conway de Macario, A.J.L. Macario); Protocol 11. Use of Fluorescent Probes for Determinative Microscopy (D.A. Stahl et al.); Protocol 12. Electron Microscopy Techniques for the Archaea (M. Firtel et al.)

Biochemistry: Protocol 13. Techniques for Anaerobic Biochemistry (K.R. Sowers); Protocol 14. Purification of Tetrahydromethanopterin-dependent Enzymes (A.A. DiMarco et al.); Protocol 15. Purification of Methylreductase and Components A2 and A3a from Methanobacterium thermoautotrophicum (C.H. Kuhner et al.); Protocol 16. Purification of Coenzymes (D.J. Stemke); Protocol 17. Purification of F420-reducing Hydrogenase from Methanobacterium formicicum (S.F. Baron, J.G. Ferry); Protocol 18. Purification of Formate Dehydrogenase from Methanobacterium formicicum (N.L. Schauer, J.G. Ferry); Protocol 19. Purification of Methanosarcina thermophila Acetate Kinase and Phosphotransacetylase Overproduced in Escherichia coli (M.T. Latimer, J.G. Ferry); Protocol 20. Purification of Carbon Monoxide Dehydrogenase from Methanosarcina thermophila (M.E. Rasche et al.); Protocol 21. Purification of Carbon Monoxide Dehydrogenase-linked Ferredoxin from Methanosarcina thermophila (A.P. Clements, J.G. Ferry); Protocol 22. Purification of Carbonic Anhydrase from Methanosarcina thermophila (B.E. Alber, J.G. Ferry); Protocol 23. Purification of Methanol-catabolizing Enzymes (J.T. Keltjens et al.); Protocol 24. Purification of Enzymes Involved in Secondary Alcohol Utilization (F. Widdel, U. Frimmer); Protocol 25. Electron Microscopy Technique for Immunocytochemical Localization of Enzymes (M. Hoppert, F. Mayer); Protocol 26. Purification of Nitrogenase (Y.-T. Chien, S.H. Zinder); Protocol 27. Preparation of Pseudomurein Endopeptidase from Methanobacterium wolfei (D. Stax et al.); Protocol 28. Preparation of Methanosarcina-disaggregating Enzyme (L. Xun et al.); Protocol 29. Purification of DNA-dependent RNA Polymerase (M. Thomm); Protocol 30. Purfication of the Histone HMf from Methanothermus fervidus (K. Sandman et al.); Protocol 31. Purification of Flagellins (K.F. Jarrell et al.); Protocol 32. Isolation and Analysis of Cell Walls (H. Konig); Protocol 33. Purification of Ether Lipids and Liposome Formation from Polar Lipid Extracts (G.D. Sprott et al.); Protocol 34. Turnover of Amino Acids Monitored by NMR Spectroscopy (M.F. Roberts); Protocol 35. Detection of Phosphorylated Small Molecules H by 31P and 13C NMR Spectroscopy (A. Gorkovenko, M.F. Roberts); Protocol 36. Extraction and Detection of Compatible Intracellular Solutes (R. Ciulla et al.)

Molecular Biology and Genetics: Protocol 37. Isolation of Chromosomal and Plasmid DNAs (K.R. Sowers); Protocol 38. Isolation of RNA (M. Thomm, H.-P. Gohl); Protocol 39. Chromosomal Mapping (J. Sitzmann, A. Klein); Protocol 40. Gene Cloning following Size Selection of Restriction Endonuclease Fragments (K. Sandman et al.); Protocol 41. Gene Selection and Cloning with Bacteriophage l Libraries (K.R. Sowers); Protocol 42. Mutagenesis of Methanococcus spp. with Ethyl Methane-sulfonate (Y.-L. Yang et al.); Protocol 43. Selection for Auxotrophic Mutants of Methanococcus (T.L. Bowen et al.); Protocol 44. Integration Vectors (A. Klein, K. Horner); Protocol 45. Transformation of Methanococcus voltae by Protoplast Regeneration (G.B. Patel et al.); Protocol 46. Transduction of Methanobacterium thermo-autotrophicum Marburg (L. Meile et al.); Protocol 47. Techniques for In Vitro Transcription Assays (M. Thomm, W. Hausner)

Appendices