Lysis of a Single Cyanobacterium for Whole Genome Amplification

• Main Authors: Richard N. Zare, Eric W. Hall, Visham Appadoo, Samuel Kim
• Format: Article
• Language: English
• Published: MDPI AG 2013-08-01
• Series: Micromachines
• Subjects: cyanobacteria; lysis; whole genome amplification; single cell; microfluidics
• Online Access: http://www.mdpi.com/2072-666X/4/3/321

Bacterial species from natural environments, exhibiting a great degree of genetic diversity that has yet to be characterized, pose a specific challenge to whole genome amplification (WGA) from single cells. A major challenge is establishing an effective, compatible, and controlled lysis protocol. We present a novel lysis protocol that can be used to extract genomic information from a single cyanobacterium of Synechocystis sp. PCC 6803 known to have multilayer cell wall structures that resist conventional lysis methods. Simple but effective strategies for releasing genomic DNA from captured cells while retaining cellular identities for single-cell analysis are presented. Successful sequencing of genetic elements from single-cell amplicons prepared by multiple displacement amplification (MDA) is demonstrated for selected genes (15 loci nearly equally spaced throughout the main chromosome).