DNA library preparation for next-generation sequencing
Library preparation is a process in which we modify DNA into a form that it is compatible for high throughput sequencing, and is becoming a key molecular biology technique. While there are an amazing variety of different library preparation methods available, I thought I'd start the the blog with a description of the classic method: - Shearing/fragmentation - End Repair - DNA clean-up - A tailing - Adaptor ligation - Size selection - Amplification - Quality control Shearing/Fragmentation The DNA needs to be in a size range that is compatible with the sequencing platform. The most commonly used sequencing platforms require DNA construct in the range of 300-500 bp, although this depends on the specific platform and the application. Fragmentation can be done by mechanical disruption through sonication like we do in our lab, but can also be done with a nebuliser or with enzymatic fragmentation . Our thoughts are that sonication/nebuliser has a lesser degree of sequence speci...