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KCL NGScourse Session1 Handout

Next generation sequencing (NGS) has advanced DNA sequencing techniques. It involves preparing a DNA library, clonally amplifying the fragments, sequencing the fragments in parallel, and analyzing the resulting data. NGS is faster, cheaper, and more accurate than first generation Sanger sequencing. It has enabled sequencing of whole genomes, including the human genome. Common NGS platforms include Illumina sequencing for short reads and Oxford Nanopore and Pacific Biosciences for long reads.

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52 views19 pages

KCL NGScourse Session1 Handout

Next generation sequencing (NGS) has advanced DNA sequencing techniques. It involves preparing a DNA library, clonally amplifying the fragments, sequencing the fragments in parallel, and analyzing the resulting data. NGS is faster, cheaper, and more accurate than first generation Sanger sequencing. It has enabled sequencing of whole genomes, including the human genome. Common NGS platforms include Illumina sequencing for short reads and Oxford Nanopore and Pacific Biosciences for long reads.

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espartawithespartilho
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Introduction to NGS

Innovation Scholar UK:


Omics Pillar
Alexandre Thiery
Eva Hamrud
Centre for Craniofacial and Regenerative Biology
What is NGS?
Learning Objectives:
• Understand the main steps of NGS sequencing from library
preparation to read processing
• Appreciate the variety of different NGS platforms available, including
the options of short or long read sequencing
DNA Sequencing

https://www.genome.gov/genetics-glossary/acgt
First generation sequencing (Sanger sequencing)

https://www.yourgenome.org/facts/timeline-history-of-genomics/
First generation sequencing (Sanger sequencing)

Add structure https://geneticeducation.co.in/what-is-the-role-of-ddntps/

https://www.sigmaaldrich.com/GB/en/technical-documents/protocol/genomics/sequencing/sanger-sequencing
First generation sequencing (Sanger sequencing)

https://www.sigmaaldrich.com/GB/en/technical-documents/protocol/genomics/sequencing/sanger-sequencing
First generation sequencing (Sanger sequencing)

https://www.sigmaaldrich.com/GB/en/technical-documents/protocol/genomics/sequencing/sanger-sequencing
Next Generation Sequencing (NGS)
…Also called massive parallel sequencing

Advantages over Sanger sequencing:


• DNA input size
• Speed
• Cost
• Accuracy

https://www.genome.gov/about-genomics/fact-sheets/DNA-Sequencing-Costs-Data
NGS Steps

1)Library preparation

2)Clonal amplification

3)Sequencing

4)Data analysis
1) Library preparation

https://emea.illumina.com/content/dam/illumina-marketing/documents/products/illumina_sequencing_introduction.pdf
2) Clonal amplification

https://emea.illumina.com/content/dam/illumina-marketing/documents/products/illumina_sequencing_introduction.pdf
2) Clonal amplification by bridge amplification

https://www.illumina.com/documents/products/techspotlights/techspotlight_sequencing.pdf
2) Clonal amplification

https://emea.illumina.com/content/dam/illumina-marketing/documents/products/illumina_sequencing_introduction.pdf
3) Sequencing

https://emea.illumina.com/content/dam/illumina-marketing/documents/products/illumina_sequencing_introduction.pdf
4) Data analysis

1) Adaptor trimming, read alignment and quality control

2) Downstream analysis – application specific


The Human Genome Project

• Occurred between 1990 - 2003


• Was mainly done using Sanger sequencing
• 3.2 billion base pairs long!
NGS platforms
Short read platforms:
• Illumnia
• Ion Torrent

Long read platforms:


• Oxford Nanopore
• Pacific Bioscience
Resources
• History of DNA sequencing: https://the-dna-universe.com/2020/11/02/a-journey-through-the-history-of-
dna-sequencing/
• Illumnia sequencing: https://www.yourgenome.org/facts/what-is-the-illumina-method-of-dna-sequencing/
• Adaptors: https://www.lubio.ch/blog/ngs-
adapters#:~:text=Adapters%20are%20key%20components%20for,combined%20with%20primers%20for%20
amplification.
• NGS overview: https://www.thermofisher.com/uk/en/home/life-science/sequencing/sequencing-learning-
center/next-generation-sequencing-information/ngs-basics/what-is-next-generation-sequencing.html
• Illumina sequencing introduction: https://emea.illumina.com/content/dam/illumina-
marketing/documents/products/illumina_sequencing_introduction.pdf
• More in depth: https://www.ebi.ac.uk/training/online/courses/functional-genomics-ii-common-
technologies-and-data-analysis-methods/next-generation-sequencing/improvements-on-the-previous-
technology/
More resources
• EBI materials on NGS: https://www.ebi.ac.uk/training/online/courses/functional-genomics-ii-common-
technologies-and-data-analysis-methods/next-generation-sequencing/
• Youtube video that explains NGS with clear animations:
https://www.youtube.com/watch?v=WKAUtJQ69n8
• Information about the Human Genome Project: https://www.genome.gov/human-genome-project

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