NGS library preparation workflow (cont.)
adapter ligation, 41, 43
adenylate 3’-ends of DNA/cDNA, 41
DNA/RNA samples, 41, 42
End Repair and size selection, 41
enrich DNA/cDNA fragments, 43
fragmentation methods, 41
PCR amplification, 41
preparation, 41
purify ligation products, 43
RNA fragments, 41
RNA-seq library preparation, 41
validate and normalize, 43
NGS technologies
Illumina sequencing technologies, 48–51
Oxford Nanopore, 52–54
PacBio, 51–52
products, 55
NGS technologiesIon Torrent, 51
NGS templates, 56
Non-coding RNAs, 144
Non-invasive prenatal testing, 32
Non-negative matrix factorization (NNMF), 23
Normalization, 165, 166
Nucleotides, 3
O
Oases, 157, 158
Oncogenes, 19, 20, 33
Organoids, 26
Oxford Nanopore Technologies (MinION), 147
advantages, 54
analyte molecule, 53
biomolecules, 54
biophysical approach, 53
Coulter’s counters, 53
ions and charged biomolecules, 53
nanoscale pores, 52
noise fluctuations, 54
PCR, 54
pioneer technology, 53
pore types, 53
sequence and epigenetic modifications, 54
toxin α-hemolysin, 53
P
Pacific Biosciences (PacBio), 51, 52, 147
Package lattice, 66
Paired-end sequencing, 115
Pan-Cancer Analysis of Whole Genomes
(PCAWG), 31
Passenger mutations, 18
Patient-derived xenografts (PDXs), 26
PCR amplification, 179
Peaks/regions, 184–186, 189
Personalized genomic medicine, 32
Personalized medicine, 2
Phasing, 94
Phenotypes, 48
Philadelphia chromosome, 21
Phred Score, 83
PolyA-capture, 41
Polymerase II (Pol II), 179
Polypeptide, 6
Post-variant calling filters
end-distance bias, 133
indel and SNP gap filters, 133
steps, 132
strand bias filter, 132
Preprocessing
NGS data, 100, 101
Proto-oncogenes, 19
Q
Quality check (QC), 147
Quality Check via FastQC program
features, FastQC, 93
overrepresented sequence, 98
per base GC content, 97
per base N content, 97
per base sequence quality, 94
per sequence GC content, 97
per sequence quality scores, 96
DNA library, 99
RNA library, 99
Per Tile Sequence Quality, 94, 96
sequence duplication levels, 98, 102
sequence length distribution, 97
sequencing reads, 92
statistics module, 93, 94
Quantitative polymerase chain reaction (qPCR),
144
R
RAS-/TP53-cancer associated mutations, 22
Reference genome, 106, 124, 152, 169, 182
Reference Genome Index
genome sequence and annotation files, 106
via Bowtie2, 108
via STAR, 106, 107
Reference-based alignment
contigs, 151
de novo strategy, 151
genome based, 150
216
Index
adapter ligation, 41, 43
adenylate 3’-ends of DNA/cDNA, 41
DNA/RNA samples, 41, 42
End Repair and size selection, 41
enrich DNA/cDNA fragments, 43
fragmentation methods, 41
PCR amplification, 41
preparation, 41
purify ligation products, 43
RNA fragments, 41
RNA-seq library preparation, 41
validate and normalize, 43
NGS technologies
Illumina sequencing technologies, 48–51
Oxford Nanopore, 52–54
PacBio, 51–52
products, 55
NGS technologiesIon Torrent, 51
NGS templates, 56
Non-coding RNAs, 144
Non-invasive prenatal testing, 32
Non-negative matrix factorization (NNMF), 23
Normalization, 165, 166
Nucleotides, 3
O
Oases, 157, 158
Oncogenes, 19, 20, 33
Organoids, 26
Oxford Nanopore Technologies (MinION), 147
advantages, 54
analyte molecule, 53
biomolecules, 54
biophysical approach, 53
Coulter’s counters, 53
ions and charged biomolecules, 53
nanoscale pores, 52
noise fluctuations, 54
PCR, 54
pioneer technology, 53
pore types, 53
sequence and epigenetic modifications, 54
toxin α-hemolysin, 53
P
Pacific Biosciences (PacBio), 51, 52, 147
Package lattice, 66
Paired-end sequencing, 115
Pan-Cancer Analysis of Whole Genomes
(PCAWG), 31
Passenger mutations, 18
Patient-derived xenografts (PDXs), 26
PCR amplification, 179
Peaks/regions, 184–186, 189
Personalized genomic medicine, 32
Personalized medicine, 2
Phasing, 94
Phenotypes, 48
Philadelphia chromosome, 21
Phred Score, 83
PolyA-capture, 41
Polymerase II (Pol II), 179
Polypeptide, 6
Post-variant calling filters
end-distance bias, 133
indel and SNP gap filters, 133
steps, 132
strand bias filter, 132
Preprocessing
NGS data, 100, 101
Proto-oncogenes, 19
Q
Quality check (QC), 147
Quality Check via FastQC program
features, FastQC, 93
overrepresented sequence, 98
per base GC content, 97
per base N content, 97
per base sequence quality, 94
per sequence GC content, 97
per sequence quality scores, 96
DNA library, 99
RNA library, 99
Per Tile Sequence Quality, 94, 96
sequence duplication levels, 98, 102
sequence length distribution, 97
sequencing reads, 92
statistics module, 93, 94
Quantitative polymerase chain reaction (qPCR),
144
R
RAS-/TP53-cancer associated mutations, 22
Reference genome, 106, 124, 152, 169, 182
Reference Genome Index
genome sequence and annotation files, 106
via Bowtie2, 108
via STAR, 106, 107
Reference-based alignment
contigs, 151
de novo strategy, 151
genome based, 150
216
Index