Next-Gen sequencing


Short Glossary of NGS terms

Amplicon Sequencing High-throughput sequencing of DNA fragments obtained by conventional PCR.
Assembly Assembly of fragment sequences into higher order structures based on their overlap and reference sequence, where appropriate.
BAM File Binary version of SAM file, a typical output of the secondary phase of data analysis.
Barcode A short unique sequence through which you can identify different samples pooled into a single library.
Bridge Amplification Amplification of fragments captured on a chip where an amplified molecule is attached to the chip by the adapter at both its ends.
Chip An instrument specific matrix where actual sequencing and detection of nucleotide incorporation takes place. Chips by different manufacturers vary significantly as to their design and capacity.
CNV Copy-Number Variations, a structural change in DNA, insertion or deletion of a longer region of DNA.
Contig The first level of the association of fragment sequences to higher structures (Fragment -> Contig).
Coverage This value indicates the coverage of an analysed sequence with respect to its length, usually expressed as a percentage; sometimes the term is also used for the depth of reading.
Data Analysis formally divided into primary, secondary and tertiary.
De-Novo Sequencing Sequencing of genetic material with no reference sequence available.
Emulsion PCR if the template has appropriate concentration, the result is of a monoclonal PCR product.
Exome A part of genome made of exons.
FASTQ A file of sequences of individual reads with corresponding quality indicators for each base. A typical result of primary analysis.
Flow Cell A type of chip used in Illumina instruments.
Fragment A short stretch of nucleic acid resulting from the fragmentation of longer stretches and sequenced. The required size of a fragment is specific to the type of experiment and sequencer possibilities.
Fragmentation Splitting of genetic material into fragments of desired sizes: mechanically (nebulisation, sonication) or enzymatically
InDel Insertion/deletion = sequencing divergences that can cause a reading frame shift.
Lane A path in a chip of Flow Cell type (Illumina).
Library A set of nucleic acid fragments which has undergone all processing steps and is ready for actual sequencing.
Long-Reads Strategy for sequencing samples prepared by Mate-Pair-End method.
Mate Pair-End-Read Strategy for sample preparation where the longer fragment (thousands of bases) is circularized using labelled adapters, the molecule is subsequently fragmented, but only the fragments containing the labelled adapters are sequenced.
Multiplex a library containing various samples labelled with bar codes.
Next-Generation Sequencing Confusing term commonly used for massively parallel sequencing technology.
Output Capacity A number of read bases in sequencing, typically measured in thousands to trillions of bases (kb, Mb, Gb, Tb), can be related to an experiment, chip, instrument, etc.
Paired-End-Read A method of reading a fragment where the fragment is first read from one end and then from the other.
Primary Analysis Conversion of raw data to sequences (ACGT), assessment of the quality of base reading, chip occupation and overall success of sequencing.
Re-Sequencing Sequencing of genetic material with reference sequence available.
Read Data output from the analysis of a single fragment (sequence).
Read Accuracy Indicates the occurrence of errors (in %) after primary analysis.
Read Depth DNA = number of times a nucleotide is read; RNA = total number of reads per sample
Read Length The number of read bases per fragment, respectively the maximum length of the fragment, which can be sequenced at a time (indicated in bases).
Run A single process cycle of the sequencer from the start-up up to obtaining raw data.
SAM File File containing alignment of fragments together with quality indicators and possibly other information.
Sample Enrichment Preparation of a sample so that it contains the maximum amount of the genetic material in question.
Scaffold The second level of the association of fragment sequences to higher structures (Fragment -> Contig -> Scaffold).
Secondary Analysis Filtration of reads on the basis of quality, alignment and assembly of reads, determination of the alignment quality indicators and assembly, administration of sequence variants.
Short-Reads Strategy for the sequencing of samples prepared using Single-End and Pair-End methods.
Shot-Gun Strategy for sample preparation where whole molecules of DNA or RNA are fragmented.
Single-Read A method of reading a fragment where the fragment is read from one end only during sequencing.
SMRT Cell A type of chip used in Pacific Biosciences instruments (Single Molecule Real Time technology).
SNP Single-Nucleotide Polymorphism = sequence divergence in the range of a single base.
SNP Calling Process of detecting SNPs in the sequences obtained.
Tertiary Analysis Contextual analysis of the results of sequencing and comparison of the results of different samples, etc.
Transcriptome Total RNA present in a cell.
Variant Calling Process of detection of sequence variants in the sequences obtained.
VCF File

A file containing information about the sequence variants identified, a typical output of the secondary phase of data analysis.


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