Life Science Research Fundamentals

The 2006 Nobel Prize in Physiology or Medicine was awarded to Andrew Z. Fire and Craig C. Mello for their discovery of RNA interference (RNAi). RNAi is a biological process in which RNA molecules can silence (inhibit) or up-or down-regulate gene expression, typically by causing the destruction of specific messenger RNA (mRNA) molecules. This collection of articles begins with an introduction to RNAi and proceeds to describe uses of this technology in various approaches to disease treatment, including gene therapy. Several laboratory protocols for silencing genes via RNAi are also provided, as are protocols for down-regulating and then rescuing those down-regulated genes, which demonstrates specificity of the approach. This eBook-a curated collection from eLS, WIREs, and Current Protocols-offers a fantastic introduction to the field of RNA interference for students or interdisciplinary collaborators.

The need for novel antibiotics is greater now than perhaps any time since the pre-antibiotic era. Indeed, the recent collapse of many pharmaceutical antibacterial groups, combined with the emergence of hypervirulent and pan-antibiotic-resistant bacteria has severely compromised infection treatment options and led to dramatic increases in the incidence and severity of bacterial infections. This collection of reviews and laboratory protocols gives the reader an introduction to the causes of antibiotic resistance, the bacterial strains that pose the largest danger to humans (i.e., streptococci, pneumococci and enterococci) and the antimicrobial agents used to combat infections with these organisms. Some new avenues that are being investigated for antibiotic development are also discussed. Such developments include the discovery of agents that inhibit bacterial RNA degradation, the bacterial ribosome, and structure-based approaches to antibiotic drug discovery. Two laboratory protocols are provided to illustrate different strategies for discovering new antibiotics. One is a bacterial growth inhibition assay to identify inhibitors of bacterial growth that specifically target conditionally essential enzymes in the pathway of interest. The other protocol is used to identify inhibitors of bacterial cell-to-cell signaling. This e-book - a curated collection from eLS, WIREs, and Current Protocols - offers a fantastic introduction to the field of antibiotics and antibiotic resistance for students or interdisciplinary collaborators.

This collection of reviews and protocols provides the reader with an introduction to the current state of knowledge on how various diseases are related to mitochondrial dysfunction. Mitochondria contain their own genome, a small, circular double-stranded DNA (mtDNA), and alterations in mtDNA may play an important role in the multistep carcinogenesis of at least some types of human cancer. In addition to mutations of mtDNA, many mitochondrial syndromes are due to abnormalities in nuclear genes related to oxidative phosphorylation (OXPHOS). Mitochondrial tRNA (MTT) gene mutations are an important cause of human morbidity and are associated with a wide range of pathology, from isolated organ?]specific diseases such as myopathy or hearing loss, to multisystem disorders with encephalopathy, gastrointestinal dysmotility, and life?]threatening cardiomyopathy. The relationship of the mitochondrion organelle to aging and longevity is also discussed. Laboratory protocols describe methodology to characterize mtDNA heteroplasmy by parallel sequencing. Each eukaryotic cell contains hundreds of mitochondria with hundreds of mitochondria genomes. Mutant and wild-type mtDNA may co-exist as heteroplasmy, and cause human disease. The purpose of this protocol is to simultaneously determine mtDNA sequence and quantify the heteroplasmic level. Another protocol describes procedures for obtaining tissue sections and cell material suitable for histological evaluation of OXPHOS activity and integrity and immunodetection of the complexes in tissue from patients suspected of mitochondrial disease. Emphasis lies on the diagnostic potential of these techniques to differentiate mtDNA from nuclear mutations. This eBook-a curated collection from eLS, WIREs, and Current Protocols-offers a fantastic introduction to the field of mitochondrial diseases for students or interdisciplinary collaborators.

With the increasing worldwide incidence of obesity and the resulting increase in metabolic disorders such as diabetes, cardiovascular diseases and some cancers, an understanding of the multitude of factors that contribute to obesity is crucial for researchers to develop more effective treatments. This collection of overviews and protocols provides the reader with an introduction to the major features of obesity research, including the interplay between energy balance, hormones and neurobiological networks. An overview is provided which describes the animal models of obesity most commonly utilized in research. A standardized laboratory protocol for producing, Diet Induced Obesity (DIO) in both mouse and rat is provided as are protocols that describe how diet formulation can be modified to generate mouse models of human metabolic pathologies. Protocols are also provided for testing the effects of dietary manipulations, caloric restriction and potential therapeutics. This eBook-a curated collection from eLS, WIREs, and Current Protocols-offers a fantastic introduction to the field of obesity research for students or interdisciplinary collaborators.

Liposomes are synthetic vesicles consisting of one or more phospholipid bilayers, able to accommodate water-and lipid-soluble molecules. They are used as a delivery system for drugs, genes and vaccines in therapeutics. This collection of reviews and laboratory protocols provides the reader with an excellent introduction to some of the many ways in which liposomes can be labeled and used in both diagnostic and therapeutic applications. Diagnostic uses include attachment of radiolabels to track nanometer-sized liposomes in the body in a quantitative fashion using PET or SPECT imaging. The ability to track the uptake of liposomes in humans and research animals on a whole-body basis provides an excellent tool for developing liposome-based drug delivery agents. The attachment of therapeutic molecules to liposomes also has great promise in cancer therapy. In addition, recent developments in the use of liposomes carrying therapeutic radionuclides for cancer therapy will be reviewed. A laboratory protocol is also provided to illustrate how liposomes are routinely used to transfect (deliver) DNA into cells. This eBook is a curated collection from eLS, WIREs, and Current Protocols on the topic of liposomes in nanomedicine. It is a great introduction to the field for students or interdisciplinary collaborators.

Epigenetics is the study of heritable changes in gene expression or cellular phenotype, caused by mechanisms other than changes in the DNA sequence. Examples include DNA methylation and histone modification. These changes may remain through cell divisions and multiple generations. Epigenetic differences among individuals may account for some of the differences between monozygotic (identical) twins. Aberrant DNA methylation is also frequently associated with human aging and diseases, such as cancer. This collection of overviews and laboratory protocols provides crucial, distilled information about the roles of epigenetics in cancer development. The overviews are geared for research scientists who need to learn more about the current understanding of epigenetic variation in humans and how the processes of DNA methylation and histone modification are regulated. The protocols give step-by-step instructions on how to detect DNA methylation using various methods such as MAPit, CHARM (arrays) and methylation-specific PCR. This eBook, a curated collection from eLS, WIREs, and Current Protocols-offers a fantastic introduction to the fields of genetics, genomics, and oncogenesis for students or interdisciplinary collaborators.

Influenza virus infections are a serious health problem. Each year, about 500 million people are infected by the virus, resulting in about 500,000 deaths worldwide. The occurrence of new influenza subtypes has caused severe pandemics, including the 2009 swine flu (vH1N1). In addition, highly pathogenic viruses, like subtypes H7N7 and H5N1, also called the fowl plague and bird flu, respectively, represent enormous economic threats to livestock farming. This book begins with descriptions of the molecular make-up of influenza viruses, their replication cycles and functions of viral proteins. A history of major influenza pandemics is provided as is a detailed article discussing how viral growth and decay is mathematically modeled to evaluate the biological parameters governing interaction between host and virus. Several laboratory protocols describe how influenza virus is handled and used in animal models to study host-pathogen interactions and test potential new therapies. This e-book-a curated collection from eLS, WIREs, and Current Protocols-offers a fantastic introduction to the field of influenza research for students or interdisciplinary collaborators.