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Single Cell Analysis Of Mrna Splicing Variants
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Book Synopsis Single-cell Analysis of MRNA Splicing Variants by : Jacob Potts
Download or read book Single-cell Analysis of MRNA Splicing Variants written by Jacob Potts and published by . This book was released on 2020 with total page 40 pages. Available in PDF, EPUB and Kindle. Book excerpt: "Single-cell RNA sequencing (scRNA-seq) and imaging technologies have revealed the transcriptomic diversity of single cells, the study of which is key to understanding biological processes and disease states. Alternative splicing of RNA contributes towards this diversity, yielding variable transcriptional profiles across different cell types within the human body. This diversity is challenging to visualize in situ because current isoform-sensitive RNA imaging methods are inefficient or are only applicable to genes with large isoform-specific exons. Here, we attempt to expand the toolbox of spatially informed RNA visualization methods by optimizing click-amplifying fluorescent in situ hybridization (clampFISH) to visualize alternatively-spliced isoforms and describe some of the challenges we encountered. We conducted a meta-analysis of combined short- and long-read sequencing data to select optimal targets for splicing analysis in human cell lines. We developed several single molecule FISH (smFISH) probe sets to evaluate efficacy of isoform-specific clampFISH probes in future experiments. Next, we modified clampFISH probes to include more landing pads (binding sites for subsequent probes) with the aim of achieving more rapid signal amplification with fewer primary probes. We observed higher target signal intensity with more landing pads, but this effect created higher off-target signal intensity as well. Finally, we tested whether adding magnesium ions to hybridization conditions would increase clampFISH probe specificity. The tested magnesium ion concentrations reduced nonspecific binding but also reduced specific binding. Our results lay the foundation for future experiments that will inform isoform distributions at a single-cell level"--Author's abstract.
Book Synopsis Interpretation, Stratification and Validation of Sequence Variants Affecting MRNA Splicing in Complete Human Genome Sequences by : Ben Chambers Shirley
Download or read book Interpretation, Stratification and Validation of Sequence Variants Affecting MRNA Splicing in Complete Human Genome Sequences written by Ben Chambers Shirley and published by . This book was released on 2013 with total page 134 pages. Available in PDF, EPUB and Kindle. Book excerpt: The Shannon Human Splicing Pipeline software has been developed to analyze variants on a genome-scale. Evidence is provided that this software predicts variants affecting mRNA splicing. Variants are examined through information-based analysis and the context of novel mutations as well as common and rare SNPs with splicing effects are displayed. Potential natural and cryptic mRNA splicing variants are identified, and inactivating mutations are distinguished from leaky mutations. Mutations and rare SNPs were predicted in genomes of three cancer cell lines (U2OS, U251 and A431), supported by expression analyses. After filtering, tractable numbers of potentially deleterious variants are predicted by the software, suitable for further laboratory investigation. In these cell lines, novel functional variants comprised 6-17 inactivating mutations, 1-5 leaky mutations and 6-13 cryptic splicing mutations. Predicted effects were validated by RNA-seq data of the three cell lines, and expression microarray analysis of SNPs in HapMap cell lines.
Author :Andras Nagy Publisher :Cold Spring Harbor, N.Y. : Cold Spring Harbor Laboratory Press ISBN 13 : Total Pages :784 pages Book Rating :4.3/5 (91 download)
Book Synopsis Manipulating the Mouse Embryo by : Andras Nagy
Download or read book Manipulating the Mouse Embryo written by Andras Nagy and published by Cold Spring Harbor, N.Y. : Cold Spring Harbor Laboratory Press. This book was released on 2003 with total page 784 pages. Available in PDF, EPUB and Kindle. Book excerpt: Provides background information and detailed protocols for developing a mouse colony and using the animals in transgenic and gene-targeting experiments. The protocols list the animals, equipment, and reagents required and step-by-step procedures. Topics include in vitro culture of preimplantation embryos, surgical procedures, the production of chimeras, and the analysis of genome alterations. The third edition adds protocols for cloning mice, modifying embryonic stem cells, intracytoplasmic sperm injection, and cryopreservation of embryos.
Book Synopsis Function and Regulation of Nucleotide Variants in RNA by : Giovanni Quinones Valdez
Download or read book Function and Regulation of Nucleotide Variants in RNA written by Giovanni Quinones Valdez and published by . This book was released on 2021 with total page 176 pages. Available in PDF, EPUB and Kindle. Book excerpt: RNA molecules harbor the information necessary for the synthesis of proteins and are essential to a wide variety of cellular processes. Variation of the RNA sequences results in significant phenotypic differences; however, the precise relationship between the two remains largely unknown. Thanks to the advent of high-throughput sequencing technologies, we now have the opportunity to study the transcriptome with unprecedented detail and characterize many different types of variants present in the RNA. In the present work, we developed novel computational approaches and performed in-depth analysis of RNA-sequencing (RNA-seq) data with the overarching goal of studying the function and regulations of nucleotide variants in RNA. We first aimed to understand the factors that regulate the most prevalent type of non-genetic nucleotide variant in human RNAs which is Adenosine-to-Inosine (A-to-I) editing. We analyzed bulk RNA-seq data obtained following the knockdown of over two hundred RNA Binding Proteins (RBPs) individually. This allowed us to study their role in the regulation of A-to-I editing at the transcriptome-wide scale. We identified several RBPs including DROSHA, ILF2/3, TROVE2, and TARDBP that significantly alter editing levels through various mechanisms including directly targeting the expression of ADAR1, protein-protein interaction, and direct binding to edited regions. Next, to study the effect of nucleotide variants, we made use of single-cell RNA-sequencing (scRNA-seq) data. This technology offers a unique glimpse of the transcriptome at the single cell-resolution. However, identification of nucleotide variants in scRNA-seq remains challenging and very few methods are available for this purpose. Here, we present scAllele, a novel method that detects both single nucleotide variants (SNVs) and microindels in scRNA-seq with high accuracy and sensitivity. In addition, scAllele identifies functional relationships between the identified variants and alternative RNA processing. We applied scAllele to scRNA-seq data derived from lung cancer patients (matched tumor and normal) and detected over 150 allele-specific splicing events that were unique to each condition or showed differential prevalence. Based on scAllele, we further developed a new method, namely T-Allele, to identify nucleotide variants and their linkage patterns in third-generation RNA-seq data. We demonstrated that the precision of variant calls by T-Allele is robust despite the relatively high sequencing error rate of this type of data. Using T-Allele, we identified up to 44 haplotype-specific alternative splicing events in each of the 8 cell lines included in our study. We also showed T-allele's ability to segregate alternative splicing events regulated genetically from those whose regulation involved other factors.
Book Synopsis Molecular Mechanism of Congenital Heart Disease and Pulmonary Hypertension by : Toshio Nakanishi
Download or read book Molecular Mechanism of Congenital Heart Disease and Pulmonary Hypertension written by Toshio Nakanishi and published by Springer Nature. This book was released on 2020-02-28 with total page 374 pages. Available in PDF, EPUB and Kindle. Book excerpt: This open access book focuses on the molecular mechanism of congenital heart disease and pulmonary hypertension, offering new insights into the development of pulmonary circulation and the ductus arteriosus. It describes in detail the molecular mechanisms involved in the development and morphogenesis of the heart, lungs and ductus arteriosus, covering a range of topics such as gene functions, growth factors, transcription factors and cellular interactions, as well as stem cell engineering technologies. The book also presents recent advances in our understanding of the molecular mechanism of lung development, pulmonary hypertension and molecular regulation of the ductus arteriosus. As such, it is an ideal resource for physicians, scientists and investigators interested in the latest findings on the origins of congenital heart disease and potential future therapies involving pulmonary circulation/hypertension and the ductus arteriosus.
Book Synopsis Molecular Biology of The Cell by : Bruce Alberts
Download or read book Molecular Biology of The Cell written by Bruce Alberts and published by . This book was released on 2002 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Book Synopsis Proceedings of the Twelfth Annual ACM-SIAM Symposium on Discrete Algorithms by : SIAM Activity Group on Discrete Mathematics
Download or read book Proceedings of the Twelfth Annual ACM-SIAM Symposium on Discrete Algorithms written by SIAM Activity Group on Discrete Mathematics and published by SIAM. This book was released on 2001-01-01 with total page 962 pages. Available in PDF, EPUB and Kindle. Book excerpt: Contains 130 papers, which were selected based on originality, technical contribution, and relevance. Although the papers were not formally refereed, every attempt was made to verify the main claims. It is expected that most will appear in more complete form in scientific journals. The proceedings also includes the paper presented by invited plenary speaker Ronald Graham, as well as a portion of the papers presented by invited plenary speakers Udi Manber and Christos Papadimitriou.
Book Synopsis Statistical Methods for Bulk and Single-cell RNA Sequencing Data by : Wei Li
Download or read book Statistical Methods for Bulk and Single-cell RNA Sequencing Data written by Wei Li and published by . This book was released on 2019 with total page 207 pages. Available in PDF, EPUB and Kindle. Book excerpt: Since the invention of next-generation RNA sequencing (RNA-seq) technologies, they have become a powerful tool to study the presence and quantity of RNA molecules in biological samples and have revolutionized transcriptomic studies on bulk tissues. Recently, the emerging single-cell RNA sequencing (scRNA-seq) technologies enable the investigation of transcriptomic landscapes at a single-cell resolution, providing a chance to characterize stochastic heterogeneity within a cell population. The analysis of bulk and single-cell RNA-seq data at four different levels (samples, genes, transcripts, and exons) involves multiple statistical and computational questions, some of which remain challenging up to date. The first part of this dissertation focuses on the statistical challenges in the transcript-level analysis of bulk RNA-seq data. The next-generation RNA-seq technologies have been widely used to assess full-length RNA isoform structure and abundance in a high-throughput manner, enabling us to better understand the alternative splicing process and transcriptional regulation mechanism. However, accurate isoform identification and quantification from RNA-seq data are challenging due to the information loss in sequencing experiments. In Chapter 2, given the fast accumulation of multiple RNA-seq datasets from the same biological condition, we develop a statistical method, MSIQ, to achieve more accurate isoform quantification by integrating multiple RNA-seq samples under a Bayesian framework. The MSIQ method aims to (1) identify a consistent group of samples with homogeneous quality and (2) improve isoform quantification accuracy by jointly modeling multiple RNA-seq samples and allowing for higher weights on the consistent group. We show that MSIQ provides a consistent estimator of isoform abundance, and we demonstrate the accuracy of MSIQ compared with alternative methods through both simulation and real data studies. In Chapter 3, we introduce a novel method, AIDE, the first approach that directly controls false isoform discoveries by implementing the statistical model selection principle. Solving the isoform discovery problem in a stepwise manner, AIDE prioritizes the annotated isoforms and precisely identifies novel isoforms whose addition significantly improves the explanation of observed RNA-seq reads. Our results demonstrate that AIDE has the highest precision compared to the state-of-the-art methods, and it is able to identify isoforms with biological functions in pathological conditions. The second part of this dissertation discusses two statistical methods to improve scRNA-seq data analysis, which is complicated by the excess missing values, the so-called dropouts due to low amounts of mRNA sequenced within individual cells. In Chapter 5, we introduce scImpute, a statistical method to accurately and robustly impute the dropouts in scRNA-seq data. The scImpute method automatically identifies likely dropouts, and only performs imputation on these values by borrowing information across similar cells. Evaluation based on both simulated and real scRNA-seq data suggests that scImpute is an effective tool to recover transcriptome dynamics masked by dropouts, enhance the clustering of cell subpopulations, and improve the accuracy of differential expression analysis. In Chapter 6, we propose a flexible and robust simulator, scDesign, to optimize the choices of sequencing depth and cell number in designing scRNA-seq experiments, so as to balance the exploration of the depth and breadth of transcriptome information. It is the first statistical framework for researchers to quantitatively assess practical scRNA-seq experimental design in the context of differential gene expression analysis. In addition to experimental design, scDesign also assists computational method development by generating high-quality synthetic scRNA-seq datasets under customized experimental settings.
Book Synopsis Alternative Splicing and Disease by : Philippe Jeanteur
Download or read book Alternative Splicing and Disease written by Philippe Jeanteur and published by Springer Science & Business Media. This book was released on 2006-10-04 with total page 265 pages. Available in PDF, EPUB and Kindle. Book excerpt: Splicing of primary RNA transcript is a quasi-systematic step of gene expression in higher organisms. This is the first book to highlight the medical implications, i.e. diseases, caused by alternative splicing. Alternative splicing not only vastly increases protein diversity but also offers numerous opportunities for aberrant splicing events with pathological consequences. The book also outlines possible targets for therapy.
Book Synopsis New Developments in Alternative Splicing Research by : Samuel DiMaggio
Download or read book New Developments in Alternative Splicing Research written by Samuel DiMaggio and published by Nova Science Publishers. This book was released on 2013 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: Alternative splicing occurs in most human genes and contributes to protein diversity by producing multiple mRNAs from each gene. In this book, the authors present new developments in alternative splicing research. Topics discussed include alternative splicing alterations in Alzheimer's disease; plant RNA-binding proteins implicate mRNA processing in abscisic acid (ABA) responses; comprehensive analyses of alternative exons in neuronally differentiated P19 cells; an epigenetic view on alternative splicing; identification of genuine alternative splicing variants for rare or long-sized transcripts; alternative RNA splicing and regulation of nitric oxide signalling; alternative splicing by analysing human mRNA diversity using data of FLJ human cDNAs; alternative splicing in human immune systems and auto-immune diseases; and poly (ADP-Ribosyl)ation regulation in alternative splicing.
Book Synopsis Alzheimer’s and Parkinson’s Diseases by : Israel Hanin
Download or read book Alzheimer’s and Parkinson’s Diseases written by Israel Hanin and published by Springer Science & Business Media. This book was released on 2013-06-29 with total page 690 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book represents the third in a series of International Conferences related to Alzheimer's (AD) and Parkinson's (PD) diseases. The first one took place in Eilat, Israel, in 1985; and the second one in Kyoto, Japan, in 1989. This book contains the full text of oral and poster presentations from the Third International Conference on Alzheimer's and Parkinson's Diseases: Recent Developments, held in Chicago, Illinois, U.S.A. on November 1-6, 1993. The Chicago Conference was attended by 270 participants. The Scientific Program was divided into nine oral sessions, a keynote presentation, and a poster session. The conference culminated in a Round Table Discussion involving all of the participants in the conference. The four and one-half day meeting served as an excellent medium for surveying the current status of clinical and preclinical developments in AD and PD. There were 59 oral presentations and 93 posters. This book incorporates a majority of both.
Book Synopsis Introduction to Single Cell Omics by : Xinghua Pan
Download or read book Introduction to Single Cell Omics written by Xinghua Pan and published by Frontiers Media SA. This book was released on 2019-09-19 with total page 129 pages. Available in PDF, EPUB and Kindle. Book excerpt: Single-cell omics is a progressing frontier that stems from the sequencing of the human genome and the development of omics technologies, particularly genomics, transcriptomics, epigenomics and proteomics, but the sensitivity is now improved to single-cell level. The new generation of methodologies, especially the next generation sequencing (NGS) technology, plays a leading role in genomics related fields; however, the conventional techniques of omics require number of cells to be large, usually on the order of millions of cells, which is hardly accessible in some cases. More importantly, harnessing the power of omics technologies and applying those at the single-cell level are crucial since every cell is specific and unique, and almost every cell population in every systems, derived in either vivo or in vitro, is heterogeneous. Deciphering the heterogeneity of the cell population hence becomes critical for recognizing the mechanism and significance of the system. However, without an extensive examination of individual cells, a massive analysis of cell population would only give an average output of the cells, but neglect the differences among cells. Single-cell omics seeks to study a number of individual cells in parallel for their different dimensions of molecular profile on genome-wide scale, providing unprecedented resolution for the interpretation of both the structure and function of an organ, tissue or other system, as well as the interaction (and communication) and dynamics of single cells or subpopulations of cells and their lineages. Importantly single-cell omics enables the identification of a minor subpopulation of cells that may play a critical role in biological process over a dominant subpolulation such as a cancer and a developing organ. It provides an ultra-sensitive tool for us to clarify specific molecular mechanisms and pathways and reveal the nature of cell heterogeneity. Besides, it also empowers the clinical investigation of patients when facing a very low quantity of cell available for analysis, such as noninvasive cancer screening with circulating tumor cells (CTC), noninvasive prenatal diagnostics (NIPD) and preimplantation genetic test (PGT) for in vitro fertilization. Single-cell omics greatly promotes the understanding of life at a more fundamental level, bring vast applications in medicine. Accordingly, single-cell omics is also called as single-cell analysis or single-cell biology. Within only a couple of years, single-cell omics, especially transcriptomic sequencing (scRNA-seq), whole genome and exome sequencing (scWGS, scWES), has become robust and broadly accessible. Besides the existing technologies, recently, multiplexing barcode design and combinatorial indexing technology, in combination with microfluidic platform exampled by Drop-seq, or even being independent of microfluidic platform but using a regular PCR-plate, enable us a greater capacity of single cell analysis, switching from one single cell to thousands of single cells in a single test. The unique molecular identifiers (UMIs) allow the amplification bias among the original molecules to be corrected faithfully, resulting in a reliable quantitative measurement of omics in single cells. Of late, a variety of single-cell epigenomics analyses are becoming sophisticated, particularly single cell chromatin accessibility (scATAC-seq) and CpG methylation profiling (scBS-seq, scRRBS-seq). High resolution single molecular Fluorescence in situ hybridization (smFISH) and its revolutionary versions (ex. seqFISH, MERFISH, and so on), in addition to the spatial transcriptome sequencing, make the native relationship of the individual cells of a tissue to be in 3D or 4D format visually and quantitatively clarified. On the other hand, CRISPR/cas9 editing-based In vivo lineage tracing methods enable dynamic profile of a whole developmental process to be accurately displayed. Multi-omics analysis facilitates the study of multi-dimensional regulation and relationship of different elements of the central dogma in a single cell, as well as permitting a clear dissection of the complicated omics heterogeneity of a system. Last but not the least, the technology, biological noise, sequence dropout, and batch effect bring a huge challenge to the bioinformatics of single cell omics. While significant progress in the data analysis has been made since then, revolutionary theory and algorithm logics for single cell omics are expected. Indeed, single-cell analysis exert considerable impacts on the fields of biological studies, particularly cancers, neuron and neural system, stem cells, embryo development and immune system; other than that, it also tremendously motivates pharmaceutic RD, clinical diagnosis and monitoring, as well as precision medicine. This book hereby summarizes the recent developments and general considerations of single-cell analysis, with a detailed presentation on selected technologies and applications. Starting with the experimental design on single-cell omics, the book then emphasizes the consideration on heterogeneity of cancer and other systems. It also gives an introduction of the basic methods and key facts for bioinformatics analysis. Secondary, this book provides a summary of two types of popular technologies, the fundamental tools on single-cell isolation, and the developments of single cell multi-omics, followed by descriptions of FISH technologies, though other popular technologies are not covered here due to the fact that they are intensively described here and there recently. Finally, the book illustrates an elastomer-based integrated fluidic circuit that allows a connection between single cell functional studies combining stimulation, response, imaging and measurement, and corresponding single cell sequencing. This is a model system for single cell functional genomics. In addition, it reports a pipeline for single-cell proteomics with an analysis of the early development of Xenopus embryo, a single-cell qRT-PCR application that defined the subpopulations related to cell cycling, and a new method for synergistic assembly of single cell genome with sequencing of amplification product by phi29 DNA polymerase. Due to the tremendous progresses of single-cell omics in recent years, the topics covered here are incomplete, but each individual topic is excellently addressed, significantly interesting and beneficial to scientists working in or affiliated with this field.
Book Synopsis Transcription and Splicing by : B. D. Hames
Download or read book Transcription and Splicing written by B. D. Hames and published by Oxford University Press, USA. This book was released on 1988 with total page 238 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book gives a co-ordinated review of our present knowledge of eukaryotic RNA synthesis.
Book Synopsis The Statistics of RNA Splicing in Single Cells by : Julia Eve Olivieri
Download or read book The Statistics of RNA Splicing in Single Cells written by Julia Eve Olivieri and published by . This book was released on 2022 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: Although the amount of single-cell RNA-sequencing (scRNA-seq) data has exponentially increased in recent years, analysis of RNA splicing in these datasets remains virtually nonexistent, largely due to the sparsity and bias of the data. In this thesis, we introduce a new method of analyzing differential splicing at the single-cell level and apply this method to make new biological discoveries. We start by introducing the SpliZ, which quantifies the alternative splicing of each gene in a single number for each cell in the dataset. We verify the validity of the SpliZ through simulation, comparison with existing methods, and re-discovery of known true positives in the human lung. Next, we apply the SpliZ to over 200,000 cells from human, mouse, and mouse lemur to create a comprehensive atlas of cell-type-resolved alternative splicing, with experimental validation of two examples. We discover that unsupervised clustering of cells based only on the SpliZ scores of RPS24 and ATP5F1C accurately recapitulates division into stromal, immune, and epithelial compartments. Correlation of the SpliZ with developmental time reveals previously unknown conserved splicing changes throughout spermatogenesis. Finally, we apply the SpliZ to spatial transcriptomics data to discover spatially-resolved RNA splicing patterns in the mouse brain, which are more significantly localized than gene expression for Myl6 and Gng13. The SpliZ opens the door to widespread analysis of alternative splicing in scRNA-seq data.
Book Synopsis Single Cell Sequencing and Systems Immunology by : Xiangdong Wang
Download or read book Single Cell Sequencing and Systems Immunology written by Xiangdong Wang and published by Springer. This book was released on 2015-03-27 with total page 184 pages. Available in PDF, EPUB and Kindle. Book excerpt: The volume focuses on the genomics, proteomics, metabolomics, and bioinformatics of a single cell, especially lymphocytes and on understanding the molecular mechanisms of systems immunology. Based on the author’s personal experience, it provides revealing insights into the potential applications, significance, workflow, comparison, future perspectives and challenges of single-cell sequencing for identifying and developing disease-specific biomarkers in order to understand the biological function, activation and dysfunction of single cells and lymphocytes and to explore their functional roles and responses to therapies. It also provides detailed information on individual subgroups of lymphocytes, including cell characters, function, surface markers, receptor function, intracellular signals and pathways, production of inflammatory mediators, nuclear receptors and factors, omics, sequencing, disease-specific biomarkers, bioinformatics, networks and dynamic networks, their role in disease and future prospects. Dr. Xiangdong Wang is a Professor of Medicine, Director of Shanghai Institute of Clinical Bioinformatics, Director of Fudan University Center for Clinical Bioinformatics, Director of the Biomedical Research Center of Zhongshan Hospital, Deputy Director of Shanghai Respiratory Research Institute, Shanghai, China.
Book Synopsis Synthetic Biology by : Christina Smolke
Download or read book Synthetic Biology written by Christina Smolke and published by John Wiley & Sons. This book was released on 2018-02-28 with total page 532 pages. Available in PDF, EPUB and Kindle. Book excerpt: A review of the interdisciplinary field of synthetic biology, from genome design to spatial engineering. Written by an international panel of experts, Synthetic Biology draws from various areas of research in biology and engineering and explores the current applications to provide an authoritative overview of this burgeoning field. The text reviews the synthesis of DNA and genome engineering and offers a discussion of the parts and devices that control protein expression and activity. The authors include information on the devices that support spatial engineering, RNA switches and explore the early applications of synthetic biology in protein synthesis, generation of pathway libraries, and immunotherapy. Filled with the most recent research, compelling discussions, and unique perspectives, Synthetic Biology offers an important resource for understanding how this new branch of science can improve on applications for industry or biological research.
Book Synopsis Nuclear pre-mRNA Processing in Plants by : A. S. N. Reddy
Download or read book Nuclear pre-mRNA Processing in Plants written by A. S. N. Reddy and published by Springer Science & Business Media. This book was released on 2008-04-16 with total page 323 pages. Available in PDF, EPUB and Kindle. Book excerpt: During the last few years, tremendous progress has been made in understanding various aspects of pre-mRNA processing. This book, with contributions from leading scientists in this area, summarizes recent advances in nuclear pre-mRNA processing in plants. It provides researchers in the field, as well as those in related areas, with an up-to-date and comprehensive, yet concise, overview of the current status and future potential of this research in understanding plant biology.
