Genome engineering / edited by Volker Gurtler, Michael Calcutt
- Bib ID:
- 10007669
- Format:
- Book
- Online Access:
- https://search.ebscohost.com/login.aspx?direct=true&scope=site&db=nlebk&db=nlabk&AN=3451958
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- Edition:
- First edition.
- Description:
-
- London, United Kingdom : Academic Press, 2023
- ©2023
- 1 online resource ( xvi, 234 pages ) : color illustrations.
- ISBN:
-
- 9780128235423 (electronic book)
- 012823542X (electronic book)
- Invalid ISBN:
-
- 9780128235409
- 0128235403
- Series:
- Methods in microbiology ; v. 52.
- Full contents:
-
- Intro
- Genome Engineering
- Copyright
- Contents
- Contributors
- Preface
- References
- Section I: Genome transformation
- Chapter 1: Genome transplantation in Mollicutes
- 1. The historical scientific context associated with genome transplantation
- 2. Mollicutes-The perfect model organisms for the establishment of GT
- 2.1. General characteristics of the Mollicutes
- 2.2. Transformation methods for Mollicutes
- 2.3. Transformable replicative plasmids
- 3. Baker's yeast-An engineering platform for microbial genomes
- 4. The GT protocol
- 4.1. Isolation of intact Mmc donor genomic DNA
- 4.2. Preparation of recipient Mcap cells
- 4.3. Transformation of Mcap with chromosomal DNA
- 4.4. Transplanted Mollicutes chromosomes
- 4.5. General comments about the GT protocol
- 4.6. Limiting factors involved in the GT process
- 4.6.1. Restriction-modification (R-M) systems
- 4.6.2. Phylogenetic distance of donor and recipient species
- 4.6.3. Selection of the recipient cell
- 4.6.4. DNA recombination
- 4.6.5. Nucleases
- 4.6.6. Toxin/antitoxin systems
- 4.6.7. Additional factors
- 5. Future perspectives
- 5.1. GT and Mollicutes
- 5.2. Adaptation of GT to bacterial species other than Mollicutes
- 6. Ethical considerations
- Acknowledgements
- Appendix
- A.1. Entrapping of intact donor chromosomes in agarose plugs
- A.1.1. Isolation of intact Mmc chromosomes from cultures
- A.1.2. Preparation of agarose plugs from yeast cultures containing modified donor chromosomes
- A.2. Genome transplantation using Mcap RE(-) as a recipient cell
- A.2.1. Release of intact chromosomes from agarose plugs
- A.2.2. Preparation of Mcap RE(-) recipient cells
- A.2.3. Transplantation of donor chromosomes into Mcap RE(-) recipient cells
- A.2.4. Screening of the transplants
- References
- Section II: Recombineering and engineering.
- 5. Learn and general considerations
- 5.1. Maximizing information from multi-factorial experiments: Sequential vs non-sequential optimisation
- 5.2. Case studies
- 6. Conclusions
- References
- Chapter 4: Recombineering
- 1. Introduction
- 2. History and development of recombineering
- 3. Molecular tools for recombineering
- 3.1. The RecBCD system
- 3.2. The RecF pathway
- 3.3. Lambda phage Red functions
- 3.4. Rac prophage encoded RecE and RecT
- 4. Steps involved in a typical recombineering experiment
- 4.1. Substrate DNA and its meticulous design
- 4.2. Provision for the Lambda Red recombination genes
- 4.2.1. For bacterial chromosomal DNA
- 4.2.2. For high and low copy number plasmids
- 4.3. Inducing the Red genes
- 4.4. Electroporation of the construct in the desired host
- 4.5. Growing and maintaining the electroporated cells
- 4.6. Selection and recombination of the clones
- 5. Uses of recombineering
- 5.1. Recombineering methods for inserting a selectable marker into the bacterial chromosome
- 5.2. Recombineering can be used for inserting non-selectable DNA fragments (Sharan et al., 2009)
- 5.2.1. Seamless method
- 5.2.2. Scarred method
- 6. Regulation and expression of recombineering gene
- 6.1. Lac promoter
- 6.2. Arabinose promoter
- 6.3. Lambda phage's own promoter-repressor system
- 7. Recombineering in various systems
- 7.1. In BAC (bacterial artificial chromosome)
- 7.1.1. Three step strategy
- 7.1.2. Four step strategy
- 7.1.3. ALFIRE (assisted large fragment insertion with red/ET recombination)
- 7.2. Recombineering in E. coli phages
- 7.3. Construction of Mycobacteriophage mutants by recombineering
- 7.3.1. Bacteriophage Recombineering of Electroporated DNA (BRED)
- 7.3.2. DADA-PCR: Deletion amplification assay PCR
- 7.3.3. BRED for point mutation
- 7.4. Recombineering in other strains.
- 7.5. Gram negative bacteria
- 7.5.1. Recombineering in Shewanella
- 7.5.2. Recombineering in Vibrio natriegens
- 7.5.3. Recombineering in Vibrio cholerae
- 7.5.4. Recombineering in Photorhabdus luminescens
- 7.5.5. Recombineering in Pseudomonas
- 7.5.6. Recombineering in Salmonella enterica
- 7.5.7. Recombineering in Klebsiella pneumoniae
- 7.5.8. Recombineering in Yersinia pestis
- 7.5.9. Recombineering in Zymomonas mobilis
- 7.6. Gram positive strains
- 7.6.1. Recombineering in mycobacteria
- 8. Future prospects of recombineering
- References
- Further reading
- Section III: CRISPR
- Chapter 5: Applications of CRISPR/Cas9 in the field of microbiology
- 1. Overview of CRISPR/Cas9 biology
- 2. Applications of CRISPR/Cas9
- 3. Recent uses of CRISPR/Cas9-based technologies in microbiology
- 3.1. Bacterial gene expression and CRISPR/Cas9
- 3.2. Bacterial resistance and CRISPR/Cas9
- 3.3. Delivery strategies via CRISPR/Cas9
- 3.4. Bacterial infections and CRISPR/Cas9
- 4. Techniques utilizing CRISPR/Cas9
- 4.1. Mouse model techniques
- 4.2. Techniques based on organoid models
- 4.3. Techniques based on cell lines
- 4.4. Techniques based on targeting miRNA
- 4.5. CRISPR/Cas9 in clinical trails
- 5. Challenges in the field of CRISPR/Cas9 system
- 6. Conclusion
- References
- Chapter 6: Genome engineering in Aspergillus niger
- 1. Introduction
- 2. Materials
- 2.1. Nucleotide preparation or construction
- 2.2. Manipulation of strains
- 2.3. Measurement of enzyme activity
- 2.4. Detection of secondary metabolism
- 3. Methods
- 3.1. Choice of appropriate Cas9 protein expression plasmids
- 3.2. Construction of sgRNAs
- 3.2.1. sgRNAs expression in vivo through plasmids
- 3.2.2. sgRNAs synthesis in vitro
- 3.3. Preparation of donor DNAs
- 3.3.1. Construction of donor DNAs with short homologous arms (39bp).
- 3.3.2. Construction of donor DNAs with long homologous arms (500-2000bp)
- 3.4. Transformation of host strains and verification of positive transformants
- 3.5. Detecting the yield of target products and evaluation of the genome edit effect or efficiency
- 3.5.1. Detecting the activity of glucose oxidase
- 3.5.2. Secondary metabolism detection
- 4. Notes
- References
- Section IV: Transformation
- Chapter 7: Natural transformation as a tool in Acinetobacter baylyi: Evolution by amplification of gene copy number
- 1. Introduction
- 2. General considerations
- 2.1. Design of the amplicon and synthetic bridging fragment
- 2.2. Chromosomal integration by natural transformation
- 2.3. Selection of amplification mutants
- 2.4. Interpreting gene copy number estimations and obtaining single-copy mutants
- 3. Material and equipment
- 3.1. Strains and culture media
- 3.2. Reagents for DNA manipulation
- 3.3. Equipment
- 4. Experimental procedures
- 4.1. Construction of the amplicon and chromosomal integration
- 4.2. Construction of the SBF and amplification of gene copy number
- 4.3. Adaptive laboratory evolution and monitoring of gene copy number over time
- 4.3.1. Adaptive laboratory evolution by serial transfer
- 4.3.2. Gene copy number analysis by quantitative PCR
- 4.4. Obtaining single-copy mutants from EASy
- 4.4.1. Isolation and screening by colony PCR
- 4.4.2. Allelic replacement in evolved populations
- 5. Summary and concluding remarks
- Acknowledgements
- References
- Chapter 8: Natural transformation as a tool in Acinetobacter baylyi: Streamlined engineering and mutational analysis
- 1. Introduction
- 2. General considerations
- 2.1. Convenience and optimization
- 2.2. Preparation of recipient cells and donor DNA
- 2.3. Introduction of DNA into cells, and growth conditions following transformation.
- Notes:
-
- Includes bibliographical references.
- Description based on online resource; title from digital title page (viewed on July 14, 2023).
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