Laboratory Methods in Microbiology and Molecular Biology

Methods in Molecular Microbiology

1st Edition - June 13, 2023
Authors: Richa Salwan, Vivek Sharma
Language: English
Paperback ISBN:
9 7 8 - 0 - 3 2 3 - 9 5 0 7 8 - 7
eBook ISBN:
9 7 8 - 0 - 3 2 3 - 9 5 0 7 9 - 4

Laboratory Methods in Microbiology and Molecular Biology describes various microbiological, biochemical, and molecular methods employed for the characterization, identific… Read more

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Laboratory Methods in Microbiology and Molecular Biology describes various microbiological, biochemical, and molecular methods employed for the characterization, identification, and analysis of actinomycetes, bacteria and fungi. The book details general guidelines, expectations, and responsibilities for good lab practices and consists of chapters that covers basic microbiological, physiological, biochemical, and molecular aspects as well as in silico analysis using various bioinformatic tools. Other topics in the book include how to make solutions, microscopy and imaging of microorganisms, sero-diagnostics, and basic concepts of phylogeny, physiology, biotechnology, soil, food, and environmental microbiology while working in laboratory. Laboratory Methods in Microbiology and Molecular Biology is an informative update to current practices and future perspectives for the field of microbial biotechnology. It aims to facilitate professors, researchers, and graduate students in monitoring the precision and accuracy of the qualitative and quantitative methods in their research.

Cover image
Title page
Table of Contents
Copyright
Contributor
Authors' biographies
Preface
Acknowledgments
Chapter 1. General guidelines, expectations, and responsibilities for inculcating good lab practices
1. Distributing/sharing lab responsibilities
2. Awareing students/researchers for research ethics/Authorship claims
3. Celebrations
4. Lab meetings
5. Reproducibility and data management
6. Scientific collection/work life balance
7. Handling reagents/Solutions and visiting fields
8. Laboratory safety
9. Personal safety
10. Preparing standard solutions
Chapter 2. Preparations of laboratory solutions
1. Guidelines for preparing reagents, buffers, etc
2. General considerations for working with RNA
3. General terms and description for the preparation of reagents
4. Selection and preparation of buffer solution
Chapter 3. Basic concepts of microbiology
1. Aim: preparation of culture media for the routine cultivation of bacteria
2. Aim: general techniques for the isolation and cultivation of bacteria
3. Aim: to preserve bacterial and fungal cultures by using different methods
Chapter 4. Microscopy and imaging analysis of microorganisms
1. Aim: to prepare wet mount slide of the given sample
2. Aim: to observe motility of bacteria using hanging drop experiment
3. Aim: preparation of bacterial smear
4. Aim: to prepare a slide for simple staining
5. Aim: to perform negative staining of bacteria
6. Aim: to prepare a slide for Gram's staining
7. Aim: capsule staining by anthony staining method
8. Aim: to perform capsule staining of the bacterial culture
9. Aim: to stain flagella of a bacterial culture
10. Aim: to perform spore staining of the given culture
11. Aim: perform spore staining using Dorner's method
12. Aim: to stain the nuclear material and study the detailed morphology of the nucleus
13. Aim: to perform lactophenol cotton blue staining of the give fungal culture
14. Aim: to perform staining of a given actinomycetes culture
15. Aim: to perform acid-fast staining of a given culture
Chapter 5. Characterization of microbes using molecular methods
1. Morphological characterization using Biolog
2. Microbial identification based on FAME
3. Electron microscopy
4. Molecular characterization of bacteria
Chapter 6. Soil microbiology
1. Aim: Isolation of Rhizobium from the given sample
2. Aim: To isolate Actinomycetes from the given soil sample
3. Aim: Isolation of Azotobacter from soil
4. Aim: To estimate soil microbial biomass by Chloroform fumigation extraction method
5. Aim: To estimate the quantity of various enzymes in soil samples
6. Aim: To study the effect of pH on the growth of microbes
7. Aim: To study the effect of temperature on growth of microbes
8. Aim: To study the effect of salt concentration on growth of microbes
9. Aim: To perform dual culture assay or direct confrontation assay
Chapter 7. Basic experiments in food microbiology
1. Aim: Evaluation of milk quality using methylene blue reduction test (MBRT) assay
2. Aim: Isolation of lactic acid bacteria
3. Aim: To prepare yogurt under laboratory conditions
4. Aim: Preparation of bread from wheat flour
5. Aim: To prepare alcohol from grape juice under laboratory conditions
6. Aim: To prepare sauerkraut from cabbage
7. Aim: To prepare cheese under laboratory conditions
Chapter 8. Basic experiments in microbial biochemistry
1. Aim: Qualitative screening of bacteria for amylase production
2. Aim: Qualitative screening of bacteria for lipase production
3. Aim: Qualitative screening of bacteria for protease production
4. Aim: Qualitative screening of bacteria for indole production
5. Aim: To perform methyl red (MR) test for detection of coliforms
6. Aim: To perform Voges-Proskauer test
7. Aim: Assessment of bacteria for utilization of sodium citrate as carbon source
8. Aim: To perform catalase test
9. Aim: Qualitative assessment of oxidase production
10. Aim: Assessment of carbohydrate fermentation
11. Aim: To perform oxidation-fermentation test
12. Aim: To perform nitrate reduction test
13. Aim: Qualitative assessment of urea hydrolysis
14. Aim: To perform triple sugar iron agar (TSIA) test
Chapter 9. Analytical techniques in microbial genomics
1. Isolation of nucleic acid (DNA and RNA) from microorganisms
2. Analysis of nucleic acid (DNA and RNA) from microorganisms
Chapter 10. Analytical techniques in molecular biology-proteomics
1. Basics of protein electrophoresis
2. SDS-PAGE
3. Consumables
4. Equipments
5. SDS-PAGE staining using Coomassie R250 or G250
6. Silver staining
7. Chemicals
8. Equipments and per-requisite
9. Method
10. Fluorescent silver staining
11. Chemicals
12. Equipments and per-requisite
13. Method
Chapter 11. Analytical techniques in proteomics
1. Aim: Isolation and characterization of extracellular protein
2. Protein dialysis and ultrafiltration
3. Protein fractionation and desalting using gel filtration chromatography
4. Use of bioinformatics in planning a protein purification
Chapter 12. Expression of recombinant proteins in prokaryotic host
1. Aim: Cloning and transformation of gene of interest in expression vector
2. Background of pET vector system and expression host
3. Equipment and reagents for cloning and expression of genes
4. Protocol
5. Optimization of insert: vector molar ratios
Chapter 13. Sero-diagnostics in microbiology
1. Enzyme-linked immunosorbent assay (ELISA) and variants of ELISA
2. Western blotting
3. Immunoprecipitation/agglutination assay using blood sample
4. To perform ouchterlony double diffusion or passive double immunodiffusion
5. To perform radial immunodiffusion assay
Chapter 14. Basic tools for next-generation DNA sequencing of bacterial genome
1. Background
2. Protocol
3. Reagents for illumina
4. Equipments
5. Protocol
6. Bioinformatic analyses
Index

Richa Salwan

Dr. Salwan is currently an Assistant Professor (Microbiology) at the College of Horticulture and Forestry (Dr. YS Parmar University of Horticulture and Forestry), Neri, Hamirpur, Himachal Pradesh, India. Dr. Salwan’s research interests and contributions are on the diversity of psychrotrophic bacteria from the Western Himalayas and their utilization for industrial applications. She has also worked on the exploration of extremophiles for industrially relevant enzymes and plant beneficial microbes for agricultural benefits. She has published two books and numerous research papers in several international journals. Dr. Salwan serves as an Academic Editor for PLOS ONE and is also a recognized reviewer for several journals including MDPI Genes, MDPI Diversity, MDPI Foods, BMC Microbiology, Journal of Plant Growth Regulation, and Microbial Ecology.

Affiliations and expertise

College of Horticulture and Forestry (Dr. YS Parmar University of Horticulture and Forestry), Neri, Hamirpur (HP), India

Vivek Sharma

Dr. Vivek Sharma is currently an Assistant Professor at the University Centre for Research and Development at Chandigarh University, Mohali (PB). He has more than 12 years of research experience exploring molecular attributes of Trichoderma. His research also involves examining the molecular aspects of microbes beneficial to plants such as Streptomyces, and Bacillus. He has published several research papers in international journals, serves as an Academic Editor for PLOS ONE, the review editor for Frontiers in Bioengineering and Biotechnology, an Associate Editor of Chemical and Biological Technologies in Agriculture, and is a member of the editorial board of Current Proteomics. He is also a recognized reviewer for journals such as the Journal of Advanced Research, Applied Microbiology and Biotechnology, Environmental Research, the Journal of Proteomics, BMC Genomics, BMC Plant Biology, AMB Express, Molecular Biotechnology, MDPI Pathogens, Folia Microbiology, Physiological and Molecular Plant Pathology and Archives of Microbiology.

Affiliations and expertise

University Centre for Research and Development, Chandigarh University, Gharuan, Mohali (PB.), India