Book

Snyder, Lori, author.

Bacterial genetics and genomics / Lori A.S. Snyder.

Boca Raton : CRC Press, Taylor and Francis, a Garland Science book, [2020]

©2020

xxiv, 389 pages : illustrations (chiefly color) ; 29 cm

"A number of genes have been implicated in the virulence of these related yet distinct pathogens, but the genes that define and differentiate the species and their behaviours have not been established. Further, a related species, Neisseria lactamica is not associated with either type of infection in normally healthy people, and lives as a harmless commensal. We have determined which of the genes so far identified in the genome sequences of the pathogens are also present in this non-pathogenic related species"-- Provided by publisher.

"Our understanding of bacterial genetics has progressed as the genomics field has advanced. Genetics and genomics complement and influence each other; they are inseparable. Under the novel insights from genetics and genomics, once-believed borders in biology start to fade: biological knowledge of the bacterial world is being viewed under a new light and concepts are being redefined. Species are difficult to delimit and relationships within and between groups of bacteria--the whole concept of a tree of life--is hotly debated when dealing with bacteria. The DNA within bacterial cells contains a variety of features and signals that influence the diversity of the microbial world. This text assumes readers have some knowledge of genetics and microbiology but acknowledges that it can be varied. Therefore, the book includes all of the information that readers need to know in order to understand the more advanced material in the book." -- Cover.

Bacterial genetics.

Bacterial genetics.

Bacterial genomes.

Bacterial genomes.

Bacteria -- Physiology.

Bacteria -- Physiology.

Includes bibliographical references and index.

Welcome to the world of bacterial genetics and genomics -- Further reading -- Part I: DNA, genes, and genomes -- 1. DNA -- Life originated form RNA with DNA evolving later -- Nucleic acids are made of nucleoside bases attached to a phosphate sugar backbone -- DNA was discovered in 1869 and identified as the genetic material 75 years later -- The first x-ray images of DNA were taken in 1937 with the structure finally solved in 1953 -- DNA consists of two bidirectional strand joined by deoxyribose sugars and nucleotide bases -- DNA is copied semi-conservatively every time a cell divides -- Replication starts at the origin of replication and requires primers -- DNA polymerase can only add bases in the 5' to 3' direction -- 2. Genes -- Genes are features in the DNA that encode proteins -- Bacterial transcription generates RNA based on the DNA sequence -- Initiation of transcription -- Transcriptional elongation -- Transcriptional termination -- Bacterial translation produces proteins based on the mRNA sequence -- Ribosomes -- Translation initiation -- Translation elongation -- Translation termination and ribosome recycling -- Coupled transcription-translation in bacteria has mRNA being made and used to produce proteins in tandem -- Bacteria can have more than one gene on an mRNA strand and form operons -- Open reading frames are regions of the DNA between termination codons -- When a CDS is a gene -- Expression of genes is controlled-not all genes are on all of the time -- Sigma factors are responsible for RNA polymerase promoter recognition -- Regulatory proteins change the level of transcription -- Repressor proteins prevent or reduce the transcription of genes -- Activator proteins contribute to the expression of genes -- Regulatory RNAs may have an impact upon transcription -- Riboswitches alter the transcript that includes their sequence -- Global regulators control the transcription of multiple genes -- Essential genes and accessory genes -- 3. Genomes -- Bacterial chromosomes carry the genetic material of the organism -- Some bacteria have multiple chromosomes -- Plasmids contribute additional genetic features -- When DNA that looks like a plasmid may actually be a chromosome -- Prophages add bacteriophage genomes to a bacterial genome -- The sizes of bacterial genomes are characteristic of bacterial species -- Contributions of the core genome to defining the species and the accessory genome to defining the strain -- Bacterial genomes are densely packed -- DNA base composition differ between coding region -- The origin of replication impact that base composition -- Genomic architecture can impact gene expression -- Conservation of the order of genetic feature between bacterial species -- Supercoiling can also influence gene expression -- distribution of noncoding genetic features in the bacterial chromosome -- Mutations in the bacterial genome -- Translocation can change the order of genetic features in a genome -- Inversion flip the DNA strand upon which genetic features are located -- recombination changes the genome -- Horizontal gene transfer introduces new genetic material -- Transformation involves bacterial uptake of DNA from its surroundings -- Conjugation is an encoded mechanism for DNA transfer from one bacterial cell to another -- The process of transduction can introduce bacteriophage DNA into a bacterial cell -- Part II: RNA, transcriptional regulation, and transcriptomes --

4. RNA -- Bacterial mRNAs are translated into proteins as they are being transcribed -- the size of mRNA is determined by the genes it encodes -- The state of the 5' end of mRNA is dictated by its promoter region -- Not all transcripts are translated into proteins -- There are untranslated regions at the 5' end of the mRNA transcript -- Features at the 3' end of the mRNA transcript can influence the expression of encoded genes -- Stability of RNA and its degradation by nucleases and hydrolysis -- Secondary structures formed by mRNAs impact ribosome binding and translation initiation -- Secondary structures formed by mRNA influence translational termination -- tertiary structure within mRNAs impact expression of the encoded gene -- RNA thermometers modify the expression of proteins from mRNA based on temperature -- Polyadenylation of mRNA is not just for eukaryotes -- Bacterial tRNAs are folded into tight structures -- tRNA transcripts undergo post-transcriptional processing -- rRna are essential components of the ribosome -- The bacterial cell also contains noncoding RNAs that can regulate other RNAs -- 5. Transcriptional regulation -- Regulation of gene expression at the level of transcription -- The classic example of transcriptional regulation: the lac operon -- The lac operon is also subject to catabolite repression -- The actions of the corepressor tryptophan on the trp operon -- An attenuation mechanism controls the expression of the trp operon -- Genes are regulated locally by trans-acting factors -- Repressors, activators, and inducers can influence the expression of many genes -- Two-component regulators sense change and alter transcription -- DNA changes in the promoter region locally regulate transcription in cis -- Programmed changes to DNA can alter transcription locally -- Sigma factors are essential for the initiation of gene transcription -- Sigma factors can orchestrate global regulation of gene transcription -- Control of sigma factor activity involves several components -- Global regulation can be influenced by the binding of chromatin proteins to the DNA -- The H-NS protein binds to DNA, making regions unavailable for transcription -- HU and IHF are homologous proteins that act in a similar way upon DNA -- The Fis nucleoid protein is involved in the regulation of rRNA transcription -- Quorum sensing causes transcriptional changes within the bacterial cell -- Biofilm formation is a specialized response to quorum sensing and other signals -- Cyclic di-GMP is involved in the regulation of a range of functions within the bacterial cell -- The small molecule ppGpp is an indicator of the state of the bacterial cell -- Protein thermosensors regulate expression of proteins via transcriptional regulation -- Stability and degradation of mRNA by ribonuclease III impacts upon whether a transcript is expressed as a protein -- Regulation of gene expression can be through the action of RNA binding proteins --

6. Transcriptomes -- The transcriptome changes over time -- The transcriptome change due to changing conditions -- Expression of genes outside the lac operon in response to glucose and lactose -- Tryptophan and its impact on the transcriptome -- Transcriptomic changes occur when bacterial cells contact host cells -- Changes in temperature can trigger changes in the transcriptome -- Expression of key proteins can indicate a response to temperature change -- Different types of thermosensors can alter gene expression due to temperature -- Global gene regulation can occur in response to iron -- Bacterial cells require nutrients and regulate gene expression to get what they need -- Each bacterial cell in a culture is different -- expression profiling provides a population level understanding of regulation -- The expression of multiple genes is coordinated together across the chromosome -- The transcriptional network landscape can have topology -- Part III: Proteins, structures, and proteomes -- 7. Proteins -- Amino acids contain an amine group, a carboxyl group, and a side chain -- the production of functional proteins from amino acids -- The inflexible nature of the peptide bond imposes limits on the amino acids -- Amino acids are generally present as zwitterions --There are 20 amino acids encoded in the standard genetic code of DNA -- The codons present in the DNA sequence in species specific -- The amino acids that can be made by bacteria are species specific -- Most amino acids are L stereoisomer amino acids -- the classification of an amino acid is determined by its side chain -- Glycine is small and flexible -- Alanine is abundant and versatile -- Arginine has a positively charged side chain -- Asparagine was the first amino acid identifies -- Aspartic acid is negatively charged and binds to positively charged molecules -- Cysteine forms disulfide bonds with other cysteines -- Glutamic acid is a large, acidic amino acid -- Glutamine has an uncharged side chain -- Histidine has a large positively charged side chain containing a ring structure -- Isoleucine has a branched side chain -- Leuicine is similar to isoleucine, although its branched side chain is configured differently -- Lysine has a long, flexible side chain -- Methionine is at the start of all translation -- Phenylalanine has a rigid ring structure side chain -- The side chain for proline loops back to the amine group -- Serine has an uncharged side chain that readily donates hydrogen -- Threonine is similar to serine with an uncharged polar side chain -- Tryptophan has a large side chain with a double ring structure -- Tyrosine has a hydrophobic ring structure side chain -- Valine has a branched hydrophobic side chain, similar to isoleucine and leucine -- Bacterial proteins can include other amino acids beyond the 20 with codons --

8. Protein folding and structure -- Primary amino acid structure is the linear sequence of amino acids joined by peptide bonds -- Secondary amino acid structure is a folding of the primary sequence of amino acids -- Tertiary amino acid structure form when secondary structure come together -- Quaternary amino acid structures form when tertiary structure come together -- Proteins are assisted in folding by chaperones -- Some proteins include more than just amino acids -- Phosphorylation adds a phosphate group to a protein, often activating the protein -- Lipids are added to proteins part-translationally, adding a hydrophobic protein -- Some proteins are modified through the addition of an oxygen -- Acetylation adds an acetyl group to a peptide chain -- Succinylation and acetylation can happen at the same time -- Methylation post-translationally adds a methyl group to a protein -- Nitrosylation of bacterial proteins can modify regulatory networks -- Modification can remove the fMet at the start of the peptide chain -- Proteins are made in the bacterial cytoplasm, but may be transported elsewhere -- Secreted proteins carry a signal to aid in their transport out of the cell -- 9. Multiprotein systems and proteomes -- Some cellular structural components are not directly encoded by genes -- Genetics of lipopolysaccharide production -- the LPS has to be assembled and translocated to the outer surface of the cell -- Peptidoglycan is built by proteins encoded in a cluster of genes -- bacterial membrane phospholipids are made by proteins -- Extracellular polysaccharides make up the bacterial capsule -- Proteins make up bacterial cell structures -- Some bacterial proteins are enzymes that actively cause change -- Bacterial secretion systems move proteins across the bacterial membranes -- The type 1 secretion system takes proteins across both membranes in one step -- Type 2 secretion systems take a protein from the periplasm out of the cell -- The type 3 secretion system can inject proteins like a syringe -- The type 4 secretion system includes conjugation systems and DNA uptake systems -- Type 5 secretion systems are proteins that secrete themselves --Type 6 secretion systems transport proteins into other cells, including other bacteria -- Gram-positive secretion systems can aid protein transport across the thick peptidoglycan layer -- Efflux pump systems transport harmful substances out of the bacterial cell -- All of the expressed proteins are the proteome -- Mass spectrometry technology enables the study of proteomes -- Proteomics aids in identification of the core genome -- Mass spectrometry is being used diagnostically to identify bacteria -- Proteomics can be used to investigate antibiotic resistance --

Part IV: genetics, genomics, and bioinformatics -- 10. Genetics -- Terms and conventions in the field of bacterial genetics are straightforward -- Humans have understood about traits and inheritance long before the term genetics -- DNA was ignored and believed to be too simple to be the genetic material of inheritance -- Bacterial genetics was the key to demonstrating the importance of DNA -- Insights following the recognition of DNA as the genetic material led us to where we are today -- Bacterial genetics is the cornerstone of all genetics -- the identification and isolation of restriction enzymes is important for genetics research -- there are four types of restriction enzymes, with type 2 being used most in laboratories -- The genetics of bacteria was unraveled using conjugation -- Physical maps can be made for any bacterial species using restriction enzymes -- Experimentation reveals whether a CDS is a gene and what its function may be -- Library generation and library screening can identify genes and their function -- Random mutagenesis identifies genes that have non-essential functions -- The functions of genes can be determined using knockout technologies -- Knockout a gene and complement it back check the phenotype is caused by the knock out gene -- Bacterial research has helped shape the field of genetics -- 11. Automation of sanger sequencing launched the era of bacterial genome sequencing -- The first genome sequence of a free-living organism was bacteria provided opportunities for new insight and innovation -- Bacterial genome sequencing required and fueled innovation -- The emergence of next-generation sequencing technologies greatly increased sequence data -- Next-generation sequencing has limitation -- Bacterial genome-sequencing projects shift focus due to nest-generation sequencing limitation -- Next generation sequencing enables a massive expansion of comparative genomics -- Nest-generation sequencing and epidemiology -- Bacterial genome-sequencing identification of the source of outbreaks -- quick, easy sequencing means bacterial genomes can be given a second look -- Bacteria genome sequencing can uncover bacteria never before studies -- Single-molecule sequencing is more sensitive and produces longer read lengths -- Two single-molecule sequencing technologies have emerged, including physical reading of the DNA -- 12. Bioinformatics -- A lot can be learned from looking at strings of A's, T's G's and C's -- Bioinformatics is essential for interpreting sequence data -- Annotation predicts features in sequence data and notes their location -- the process of creating an annotation starts with the DNA sequence data -- Multiple line of investigation into the sequence data feature support the annotation -- Annotations tend to start with potential genes -- Annotations tend to start with potential genes -- Homology and conserved protein domains can help identify the potential function of a CDS -- Automated annotations rapidly produce an annotation that needs manual curation -- Some features in sequence data and annotation data can confuse new annotations -- Naming genes is not straightforward, with some genes having more than one name -- Annotation errors, including spelling mistakes, can spread from one annotation to many others -- Gene locus identifiers are handy for labeling features in annotations, but reveal nothing about function -- there are three major public databases for sequence data -- Comparative genomics find that there are commonly shared genes and unique genes -- Comparative genomics can be done without assembly or annotation of sequencing data -- Horizontally transferred gene sequences tend to carry a signature that can identify them -- Genome sequence analysis can find unexpected features in the sequence data -- Comparative genomics on closely related strains can reveal biologically important information -- Comparative genomics between non-related species gives insight into bacterial evolut ion -- Strain identification using sequencing data is a powerful tool for tracking bacterial transmission -- Function predictions can be made based on sequence similarity -- Part V: Bacterial response, adaptation, and evolution --

13. Bacterial response -- Studying responses often happens in pure bacterial cultures -- Two-component regulatory systems enable bacteria to respond to their environment -- Bacteria decrease host glucose levels to impair the hos immune response -- Bacteria modulate the immune response using the type 3 secretion system -- Some bacteria cheat and let others do all the work their type 3 secretion systems -- A response might only be appropriate when the population is large -- Quorum sensing makes a beautiful bio luminescent glow in the ocean and in the lab -- Quorum sensing is a process to tally the bacterial cells -- Biofilms mature due to quorum sensing signals -- Quorum sensing has cheaters -- Going from free living to biofilm involves changes in gene expression -- Biofilm dispersal is regulated by different elements between different bacterial species-- c-di-GMP plays a key role in biofilm regulation in P. aeruginosa -- Bacteria have their own immune system to protect them from bacteriophages -- 14. Bacterial adaptation -- Within a niche, bacteria have to adapt to their peers and other bacteria -- GlcNAc has a role as a signaling molecule as well as being part of the bacterial cell wall -- Competitor bacteria can be killed with specialized type 6 secretion systems -- Caulobacter differentiate between motile and sessile cells -- Staphylococcus aureus secrete several proteins to inhibit host defenses as part of adapting their niche to their needs -- Intracellular bacteria adapt to life inside the cells of the host -- Mycobacterium tuberculosis adapts both itself and its host -- Legionella adapt by knowing when not to grow -- Group A streptococci within the host experience adaptation, mutation, and death -- Adaptation of the host to enhance spread of the infection -- Listeria monocytogenes can adapt to an intracellular or soil niche -- Environmental bacteria like lactobacillus plantarum can live in a wide variety of niches -- Pseudomonas aeruginosa adapts to live in a wide variety of environments -- Mastitis-causing bacteria streptococcus uberis can adapt to different niches within cows -- Bacteria adapt to avoid recognition by the host immune system through antigenic variation -- Several different species use gene conversion as a mechanism of antigenic variation -- Phase variation is an important means of adaptation, but is not a means of response -- Small noncoding RNAs also have a role in enabling bacteria to adapt -- 15. Evolution can be studied within bacterial cultures -- bacteria can evolve within the host and we can see this happen with sequencing technologies -- Antibiotic resistance is an easily observable evolutionary event -- Mutations can be introduced into bacterial DNA by a variety of factors -- Yersinia pestis, causing plague, has evolved from Yersinia pseudotuberculosis -- Neisseria meningitidis, causing meningococcal meningitis and septicemia, acquired its capsule fairly recently -- The number for pseudogenes in a species can reveal how recently it has adapted to a new niche -- Evolution of the bacterial surface to cope with the immune system and vaccines -- Horizontal gene transfer can bring new genes into a species, contributing to its evolution -- the particular nature of an environmental niche can create opportunities for evolution -- It is possible for completely new genes to evolve -- VI: Gene analysis, genome analysis, and laboratory techniques --

16. Gene analysis techniques -- Sequence searches are done to find out what else in similar to this gene -- Before there was BLAST, there was FASTA -- BLAST quickly finds the most similar sequences -- There are five basic versions of BLAST, addressing different search tasks -- There are other version of BLAST that do specialist searches -- Searches can look for more than just similarities -- Alignments of similar sequences are useful for further analysis -- Local alignments to compare the portions of the sequence that are similar -- Global alignments will align any sequences, similar or not -- More complex comparisons need multiple sequence alignment algorithms -- Protein localization can be predicted from the amino acids -- DNA sequence to gene to amino acid sequence to 3D protein structure, ideally -- De novo protein structure predictions base structures just on the amino acids -- Transmembrane helix prediction can find membrane proteins -- Homology modeling of proteins bases structure on known structures -- Protein threading can suggest a protein structure based on protein fold similarity -- Some gene tools are used to help designing laboratory experiments -- 17. Genome analysis techniques -- A few things happen to the genome sequencing data before the search for genes -- Identification of features in genomic data is a key aspect of analysis -- Automated annotation pipelines usefully combine feature identification tools -- visualization of an automatically generated annotation can aid manual curation -- Comparison show orthologues and paralogues, revealing evolutionary relationships between genes -- Genomes can be aligned, just like genes can be aligned -- Mauve genome alignments make stunning figures, as well as being a useful research tool -- there is value in typing data, even in the genomics age -- Galaxy provides a full analysis suite for biological data --

18. Laboratory techniques -- The study of bacterial genetics and genomics -- DNA extraction using phenol produces pure, large qualities of DNA -- Phase separation and DNA precipitation in phenol DNA extraction result in isolated DNA -- Additional considerations for phenol DNA extraction can improve the outcome -- Most DNA extractions use columns -- Troubleshooting DNA extraction can increase yield and quality of the DNA -- A quick (and dirty) DNA extraction can be achieved by boiling -- The first recombination DNA experiments in the 1970s were made possible because of restriction enzymes, which are still used today -- Set up a restriction digestion with the optimal reaction conditions -- There are a few additional considerations to remember when doing restriction digestions -- Restriction digestions are used to change DNA sequences and join sequences together -- Cut ends DNA need to be ligated together to complete cloning -- Important considerations when performing ligation -- Cloning of sequences is often important in bacterial genetics and genomic research -- TA cloning exploits a feature of PCR to rapidly clone sequences -- Some commercially available kits augment ligation and cloning with accessory proteins and exploitation of other systems -- Antibiotic resistance markers on plasmids help us find the transformed bacterial colonies -- Blue-white screening helps us find the colonies transformed with plasmids with the insert -- Laboratory techniques of molecular biology are able to copy segments of DNA in processes similar to replication -- PCR can be altered slightly to address experimental needs -- Site-directed mutagenesis systems help researchers make specific changes to DNA -- Loop-mediated isothermal amplification (LAMP) quickly amplifies DNA at a single temperature -- Following in vitro manipulation of DNA, it has to be transformed into a bacterial cell -- Calcium chloride provides a quick method to obtain competent cells for immediate use -- Chemically competent cells with Inoue buffer have the best reputation for good rates to transformation and reliability -- Chemically competent cells can be made with TSS buffer -- Transformations using chemically competent cells use similar methods, regardless of how the cells were made -- Electroporation provides an alternative to chemically competent cells -- The process of electroporation is sensitive to salts, but quick to perform -- Expression studies rely on extraction of high-quality RNA, which means controlling RNases -- RNA extraction columns work similarly to DNA extraction columns work similarly to DNA extraction columns, with some slight variations -- Acidic phenol extraction of RNA makes high-quality, pure RNA -- Part VII: Applications of bacterial genetics and genomics -- 19. Biotechnology -- Biotechnology is far older than genetic engineering -- Biotechnology impacts many aspects of our lives and of research -- Large quantities of bacteria are grown in bioreactors to yield large quantities of recombinant proteins -- Human insulin expressed in Escherichia coli is a classic example of biotechnology -- Many recombinant drugs have been made since insulin -- Recombinant production of influenza virus vaccines -- Live recombinant vaccines use live bacteria to deliver antigens -- Bioremediation uses the microbial world to correct the pollutants we have introduced into the natural world -- Bioremediation using bacteria present in the environment can help us reclaim sites -- Genetic modification for bioremediation can provide organisms with new features -- Bacteria can be renewable source of bioenergy -- 20. Infectious diseases -- The study of bacterial pathogen genes has led to new drugs to control infectious diseases -- Genomics can aid in the search for new antibiotics -- Some old drugs are getting a new lease of life due to greater depth of understanding -- Bacterial genomics has led to the development of new vaccines -- Reverse vaccinology is providing leads fo r several bacterial diseases -- New drugs are being developed that will contain the virulence of bacteria -- Monoclonal antibody therapy is useful for a variety of human diseases including infectious diseases -- Sequencing changes our understanding of the virulence factors that are important -- Gene sequencing and genome sequencing improves the resolution of epidemiology of bacterial infectious diseases -- Genome sequencing can improve infection control for surgical site infections -- Horizontal gene transfer between pathogens revealed by sequencing is improving our understanding of infections that could impact transplant recovery -- 21. Bacteriophages -- Bacteriophages have been studied for just over 100 years -- Bacteriophages cannot replicate without bacterial cells -- Some bacteriophages enter latency for a period before replication -- The MS2 bacteriophage has a very small genome and was the first genome sequenced -- Important discoveries about genetics have been made by studying bacteriophage -- The T4 bacteriophage has a characteristic morphology -- The X174 bacteriophage was the first DNA genome sequenced -- transduction is an important source of horizontal gene transfer for bacteria -- Bacteriophages also contribute to bacterial evolution through chromosomes rearrangements -- Bacteriophage and prophage genome evolution can provide interesting insights -- Bacteria have evolved strategies to avoid bacteriophage infection -- Even if bacteria become infected by bacteriophage nucleic acids, they can still fight back -- Bacteriophage resistance that fights back and uses the bacteriophages for its own ends -- Bacteriophage therapy is a potential alternative treatment for antimicrobial resistance bacteria.

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