Chapter title |
Emulsion PCR: Techniques and Applications
|
---|---|
Chapter number | 4 |
Book title |
Clinical Applications of PCR
|
Published in |
Methods in molecular biology, February 2016
|
DOI | 10.1007/978-1-4939-3360-0_4 |
Pubmed ID | |
Book ISBNs |
978-1-4939-3358-7, 978-1-4939-3360-0
|
Authors |
Kanagal-Shamanna, Rashmi, Rashmi Kanagal-Shamanna M.D., Rashmi Kanagal-Shamanna |
Editors |
Rajyalakshmi Luthra, Rajesh R. Singh, Keyur P. Patel |
Abstract |
Emulsion PCR (EmPCR) is a commonly employed method for template amplification in multiple NGS-based sequencing platforms. The basic principle of emPCR is dilution and compartmentalization of template molecules in water droplets in a water-in-oil emulsion. Ideally, the dilution is to a degree where each droplet contains a single template molecule and functions as a micro-PCR reactor. Here, we discuss the basic principles, advantages, and challenges of applications of emPCR in clinical testing. We describe the methods of preparation and enrichment of template-positive Ion PGM™ Template OT2 200 Ion Sphere™ Particles (ISPs) on the Ion Personal Genome Machine(®) (PGM™) System. For routine clinical testing, following library generation, we employ the automated Ion OneTouch™ System that includes the Ion OneTouch™ 2 and the Ion OneTouch™ ES instruments for template generation and enrichment of template-positive ISPs, respectively. |
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