Microrna Expression Detection Methods

MicroRNAs (miRNAs), endogenous noncoding regulatory mRNAs of - nucleotides, have rapidly emerged as the central players in gene expression regulation. Owing to their ever-increasing implications in the control of various biological and pathological processes, miRNAs have now been considered novel biomarkers of various human diseases including, cancer, viral disease, cardiov- cular disorders, metabolic disturbances, etc. Particular expression pro?les have been associated with particular pathological states. Expression pro?ling of miR- NAs have therefore become extremely important not only for fundamentalists but also for clinicians. However, the methodologies used for detecting protein-coding mRNAs cannot be directly applied to miRNAs because of their small size. Over the past years, researchers have made great efforts to developing techniques suitable for miRNA detection and quanti?cation; a wide spectrum of creative and inno- tive techniques (more than 30 different methods) have been invented and validated. It has come to the time now to summarize these methods and present them in an orderly manner for better understanding and utilization of these methods to miRNA research and applications. In particular, the development of methods for quantifying circulating miRNAs opens up a fascinating opportunity for realizing miRNA as diagnostic and prognostic biomarkers of human disease. A book on this subject may help boosting up the passion of researchers to further improve the existing techniques and develop more new methods to ?t to new application needs. These considerations prompted us and urged us to undertake the work: writing a book focusing on miRNA expression detection methods.

This updated volume reflects new and evolved techniques to study detection, profiling, and manipulation of microRNAs (miRNAs) in plants and animals. After overviews of how best to detect, identify, and validate microRNAs, the book continues by exploring state-of-the-art protocols for microRNA detection, approaches to profile the expression level of microRNAs, spatial expression analysis, describe in silico analysis of microRNAs and their targets, as well as protocols for functional analysis of microRNAs and their targets by CRISPR/Cas. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step and readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and up-to-date, MicroRNA Detection and Target Identification: Methods and Protocols, Second Edition aims to ensure successful results in the further study of this vital field.

This book summarizes microRNA (miRNA) biology in a variety of pathological processes, emphasizing the significant potential applications of miRNA in diagnostics and prognostics, as well as novel drug targets. The conventional techniques used for miRNA detection including standard PCR, Northern blotting, microarray and clone methods are addressed. Recent emerging strategies in miRNA detection and quantification with superior flexibility and adaptability, such as novel molecular biological techniques and locked nucleic acid (LNA) modified probes, as well as nanotechnology-based approaches, are also included. The book also highlights the latest advances in clinical-related miRNA detection methods in living cells, circulating blood and tissue, such as in situ hybridization (ISH) and molecular imaging techniques, which are useful to elucidate the biogenesis and biological function of miRNAs in vivo. Finally, the respective advantages and drawbacks of various detection techniques in this fast-moving field are discussed, along with the challenges and promising new directions. This book offers a valuable resource for analytical chemists, biologists and physicians involved in miRNA research. Dr. Xueji Zhang and Dr. Haifeng Dong are Professors at the School of Chemistry & Biological Engineering, University of Science & Technology Beijing (USTB), China. Dr. Yaping Tian is a Professor at the Department of Clinical Biochemistry, Chinese PLA General Hospital and Military Medical School, China.

MicroRNAs (miRNA) are tiny bits of genetic material that were unknown nearly 10 years ago but now represent an exciting field of study in biology. Upon their discovery, researchers revealed for the first time a new mechanism by which microRNA can stop the function of messenger RNA (mRNA) by literally cutting it in half, interfering with the normal function of specific messenger RNAs in gene expression. This "expression" of genes that code for essential proteins is essentially what controls whether a cell turns into a liver, lung, or brain cell, for example. Understanding what activates this process – or stops it – is a key to understanding the biological process and builds a foundation for advances in medicine and other fields. This volume in Methods in Enzymology presents valuable methods for studying MicroRNA, with three sections covering identification of MicroRNAs and their targets; MicroRNA expression, maturation and functional analysis; and MicroRNAs and disease.

This second edition provides updated and comprehensive methods on miRNA biogenesis and their role in the development and progression of various human diseases. Chapters detail miRNA biogenesis, isolating RNA, extracellular vesicles (EVs), circulating miRNAs, analyzing miRNA and miRDeep-P2, protocols for total RNA isolation from cells, cell-derived products, isolation and characterization of exosomes, serum, plasma specimens, and software tools. Written in the successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible protocols, and notes on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, MicroRNA Profiling: Methods and Protocols, Second Edition aims to provide comprehensive and accessible methods to undergraduate, graduate, and established scientist.

In miRNomics: MicroRNA Biology and Computational Analysis, expert researchers in the field present an overview of the current state of the art and aim to put the respective areas of research into a larger perspective. These include methods and techniques ranging from miRNA biogenesis, their biological function, computational analyses to their medical implications and applications. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and key tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, miRNomics: MicroRNA Biology and Computational Analysis seeks to aid scientists in the further study into miRNA research and statistics.

Focused manuscript on the potential use/role of miRNAs in bioprocessing, specifically the production of complex proteins in mammalian cells. With that in mind I propose a draft list of topics/chapters along the following lines: Intro on CHO/bioprocessing/engineering challenges to set scene, Genomic organization, biogenesis and mode of action, Identifying miRNA targets: Computational prediction, transcriptomics, proteomices, UTR analysis, etc., miRNA expression in Chinese Hamster Ovary cells, miRNAs as engineering targets: pathway manipulation to impact bioprocess phenotypes, miRNAs as biomarkers, Detection methods: Northern, PCR, hybridization arrays, Next Gen Seq, Manipulation of expression in cultured cells: Transient/stable disregulation, Knockout.