This book addresses the various aspects of the RNA-seq technique, especially its application in drug discovery and development, including the identification of new drug targets, the prediction of drug activity or interactions, personalized medicine and toxicogenomics.
Inhaltsverzeichnis
Chapter 1: Introduction to RNA-Sequencing and Quality Control
1. 1: What is RNA-Sequencing?
1. 2. Overview of RNA-sequencing
1. 2. 1. Isolation of the RNA and RNA quality check
1. 2. 2 Selection and depletion of particular RNA
1. 2. 3 Fragmentation
1. 2. 4 Reverse transcription to generate cDNA and adaptor sequences
1. 2. 5 Single-end and Pair-end Sequencing
1. 3. RNA-seq Sequencing techniques:
1. 3. 1 Roche 454
1. 3. 2 Illumina platform
1. 3. 3 Small-scale RNA-seq platform
1. 4. RNA-seq file format
1. 4. 1 Output file: Fastq file
1. 4. 2 Mapped file: SAM/BAM/BIGWIG formats
1. 4. 3 GTF file
1. 4. 4 BED File
1. 5. Quality control of RNA-seq Data
1. 5. 1 Basic Usage of the Public Server Galaxy
1. 5. 2 FastQC program
1. 6. Advantages of RNA-seq over microarrays
1. 7. Summary
Chapter 2: Read Alignment and Transcriptome Assembly
2. Introduction
2. 1 Transcriptome Assembly Methodology
2. 1. 1 De novo assembly
2. 2 Genome-guided Assembly
2. 2. 1 Unspliced Aligners: Burrows-Wheeler Transform (BWT)
2. 2. 2 Unspliced Aligners: Seed Methods
2. 3 Spliced Aligners
2. 3. 1 TopHat
2. 3. 2 HISAT2
2. 3. 3 STAR
2. 4 Summary
Chapter 3: Normalization and Downstream Analyses
3. 1 Introduction
3. 2 Quantification of Transcript Abundance
3. 3 Raw Counts Extraction
3. 3. 1 Rsubread and featurecounts
3. 3. 2 Normalization Methods
3. 4 Differential gene expression analysis
3. 4. 1 DESeq2
3. 4. 2 EdgeR
3. 4. 3 Ballgown
3. 5. Visualization of differential expression
3. 5. 1 Integrative Genomics Viewer (IGV)
3. 5. 2 UCSC Genome Browser
3. 5. 3 Heatmaps
3. 5. 4 Volcano Plots
3. 5. 5 DAVID Pathway Analysis
3. 6. Summary
Chapter 4: Constitutive and Alternative Splicing Events
4. 1. Introduction to RNA splicing
4. 1. 1 What is splicing?
4. 1. 2 Molecular mechanism of splicing
4. 1. 3 Alternative Splicing
4. 2. Differential splicing analysis
4. 2. 1 Cuffdiff 2
4. 2. 2 DiffSplice
4. 2. 3 DEXSeq
4. 2. 4 edgeR
4. 2. 5 LIMMA
4. 3. Summary
Chapter 5: The role of transcriptomics in identifying fusion genes and chimeric RNAs in cancer
5. 1 Introduction to fusion genes
5. 1. 1 What is a fusion gene?
5. 1. 2 Mechanisms that generate new fusion genes
5. 1. 3 Fusion RNA transcripts
5. 1. 4 The connection between fusion genes and non-coding RNAs
5. 2 Detection methods for identification of fusion genes and chimeric proteins
5. 2. 1 Guided detection approaches
5. 2. 2 High-throughput sequencing-based detection methods
5. 3 Summary
Chapter 6: MiRNA and RNA-seq
6. 1 Non-coding RNAs
6. 2 MiRNAs
6. 3 LncRNAs
6. 4 miRDeep2
6. 5 Applications
6. 6 Summary
Chapter 7: Toxicogenomics and RNA-seq
7. 1 Introduction of toxicity
7. 1. 1 Traditional toxicity study
7. 2 Toxicogenomics
7. 2. 1 What is Toxicogenomics?
7. 2. 2 Advantages: Mechanisms and predictive toxicology
7. 2. 3 Mechanisms of toxicity:
7. 2. 4 Limitations of toxicogenomics:
7. 3 Methods for toxicogenomics data analysis
7. 3. 1 Identification of Differentially expressed genes
7. 3. 2 Signature matching
7. 3. 3 Gene Networks
7. 3. 4 Co-expression networks
7. 4 Toxicogenomics databases
7. 4. 1 Comparative Toxicogenomics Database (CTD)
7. 4. 2 Japanese Toxicogenomics Project (TGP)
7. 4. 3 DrugMatrix
7. 5 Comparing microarray vs. RNA-seq
7. 6 Summary
Chapter 8: Drug Discovery and Traditional Medicine
8. 1 Introduction
8. 2 What is herbal medicine?
8. 2. 1 Traditional medicine
8. 2. 2 Herbal medicine
8. 2. 3 Use of database for bioactive compound example
8. 2. 4 Properties of candidate herbal compounds
8. 2. 5 RNA-seq and herbal medicine
8. 3 Mining functional genes of medicinal plants
8. 3. 1 Mining functional genes of medicinal plants
8. 3. 2 Discovery of secondary metabolites and their metabolic pathways
8. 3. 3 Discovery of developmental mechanisms
8. 3. 4 Development of molecular markers to improve plant breeding
8. 3. 5 Identification of target genes and molecular mechanisms of herbal drugs
8. 3. 6 Synergism of herbal compounds in pathway regulation
8. 3. 7 Herbal medicine toxicity
8. 3. 8 Natural drug repurposing
8. 4 Summary
Chapter 9: Single-Cell RNA-sequencing
9. Introduction to single-cell sequencing
9. 1 Microdroplet approaches to cell capture
9. 2 Non-microfluidic approaches to cell capture
9. 3 Single-cell processing: Cell Ranger
9. 4 STARsolo
9. 5 DropletUtils
9. 6 Seurat
9. 7 Limitations
9. 8 Applications
9. 9 Summary
