graph, 152
intronic genomes, 152
overlapping reads, 152
RNA-Seq data, 152
sequencing reads, 152
splice-aware aligners, 152
STAR, 154–156
TopHat2, 152–155
transcriptome, 150–152
Reference-based RNA-Seq analysis, 148
Reference-genome based alignment, 150
Reference-transcriptome based alignment, 150, 152
Ribonucleic acid (RNA)
code, 5
intronic structures, 5
mRNA, 2
transcriptomics, 5
types, 5
Ribosomal RNA (rRNA), 146, 147
RNA extraction, 169
RNA integrity numbers (RIN), 146
RNA molecules, 144
RNA quality, 146
RNA sequencing methods collection, 11
RNA-Seq analysis
cDNA, 145
DE analysis, 166, 167
de novo assembly (see De novo assembly)
downstream analysis, 145, 146
enzymatic fragmentation, mRNA molecules,
146
functional analysis, 168
functional annotation, de novo transcripts, 160
gene expression quantification (see Gene
expression quantification)
miRNA, 146
NGS technology, 146
non-coding RNAs, 146
normalization, 165, 166
poly(A) capture, 146
post-alignment/assembly assessment, 160, 161
protocol, 146
QC, 147, 148
reference-based alignment (see Reference-based
alignment)
RNA quality, 146
RNA-Seq library preparation, 147
sequence pre-process, 147, 148
sequencing platforms, 147, 148
siRNA, 146
statistics, 160, 161
visualization, mapped reads, 161
workflow, 146
RNA-Seq library preparation, 41, 44, 147
RNA-Seq reads, 119
RNA-sequencing, 116
RStudio, 66
S
SAM (Sequence Alignment/Map) format
alignment information, 84
CIGAR, 84–86
description of TAGs, 85
format specification, 84
into BAM, 87
TAB-delimited text format, 83
SAMtools, 160
Sequence alignment
alignment definition, 113
downstream analysis, NGS data, 112
in molecular biology, 113
software tools, 113
unit edit distance (edist), 113
Sequence databases, 113
Sequence Read Archive (SRA), 92, 148
Sequencing by synthesis (SBS), 48, 49
Sequencing coverage, 80, 81
Sequencing read, 82
Silencer, 9
Single base substitutions, 131
Single-cell DNA/RNA sequencing
cancer, 33
cellular program, 27
challenges, 27
methodologies, 27
microfluidics and nanotechnology, 27
subclonal tumors, 27
technological advances, 27
Single-cell RNA-Seq (scRNA-Seq), 147
Single-cell sequencing, 40
Single-molecule, real-time (SMRT), 51
Small interfering RNA (siRNA), 146
Smith–Waterman algorithm, 115, 116
SMRT sequencing technology, 51
SMRTbell library, 51
SNVs (single nucleotide variants), 131
Solid-phase amplification, 49
Somatic gain-of-function mutations, 19
Somatic mutations, 18, 19
Splice-aware aligners, 116, 150
Spliced Transcripts Alignment to a Reference
(STAR), 117, 150, 154
advantages, 117
Index
217
intronic genomes, 152
overlapping reads, 152
RNA-Seq data, 152
sequencing reads, 152
splice-aware aligners, 152
STAR, 154–156
TopHat2, 152–155
transcriptome, 150–152
Reference-based RNA-Seq analysis, 148
Reference-genome based alignment, 150
Reference-transcriptome based alignment, 150, 152
Ribonucleic acid (RNA)
code, 5
intronic structures, 5
mRNA, 2
transcriptomics, 5
types, 5
Ribosomal RNA (rRNA), 146, 147
RNA extraction, 169
RNA integrity numbers (RIN), 146
RNA molecules, 144
RNA quality, 146
RNA sequencing methods collection, 11
RNA-Seq analysis
cDNA, 145
DE analysis, 166, 167
de novo assembly (see De novo assembly)
downstream analysis, 145, 146
enzymatic fragmentation, mRNA molecules,
146
functional analysis, 168
functional annotation, de novo transcripts, 160
gene expression quantification (see Gene
expression quantification)
miRNA, 146
NGS technology, 146
non-coding RNAs, 146
normalization, 165, 166
poly(A) capture, 146
post-alignment/assembly assessment, 160, 161
protocol, 146
QC, 147, 148
reference-based alignment (see Reference-based
alignment)
RNA quality, 146
RNA-Seq library preparation, 147
sequence pre-process, 147, 148
sequencing platforms, 147, 148
siRNA, 146
statistics, 160, 161
visualization, mapped reads, 161
workflow, 146
RNA-Seq library preparation, 41, 44, 147
RNA-Seq reads, 119
RNA-sequencing, 116
RStudio, 66
S
SAM (Sequence Alignment/Map) format
alignment information, 84
CIGAR, 84–86
description of TAGs, 85
format specification, 84
into BAM, 87
TAB-delimited text format, 83
SAMtools, 160
Sequence alignment
alignment definition, 113
downstream analysis, NGS data, 112
in molecular biology, 113
software tools, 113
unit edit distance (edist), 113
Sequence databases, 113
Sequence Read Archive (SRA), 92, 148
Sequencing by synthesis (SBS), 48, 49
Sequencing coverage, 80, 81
Sequencing read, 82
Silencer, 9
Single base substitutions, 131
Single-cell DNA/RNA sequencing
cancer, 33
cellular program, 27
challenges, 27
methodologies, 27
microfluidics and nanotechnology, 27
subclonal tumors, 27
technological advances, 27
Single-cell RNA-Seq (scRNA-Seq), 147
Single-cell sequencing, 40
Single-molecule, real-time (SMRT), 51
Small interfering RNA (siRNA), 146
Smith–Waterman algorithm, 115, 116
SMRT sequencing technology, 51
SMRTbell library, 51
SNVs (single nucleotide variants), 131
Solid-phase amplification, 49
Somatic gain-of-function mutations, 19
Somatic mutations, 18, 19
Splice-aware aligners, 116, 150
Spliced Transcripts Alignment to a Reference
(STAR), 117, 150, 154
advantages, 117
Index
217