A Colony Multiplex Quantitative PCR-Based 3S3DBC Method and Variations of It for Screening DNA Libraries: e0116997

A DNA library is a collection of DNA fragments cloned into vectors and stored individually in host cells, and is a valuable resource for molecular cloning, gene physical mapping, and genome sequencing projects. To take the best advantage of a DNA library, a good screening method is needed. After des...

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Bibliographic Details
Published in:PloS one 2015-02, Vol.10 (2)
Main Authors: An, Yang, Toyoda, Atsushi, Zhao, Chen, Fujiyama, Asao, Agata, Kiyokazu
Format: Article
Language:English
Online Access:Get full text
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Summary:A DNA library is a collection of DNA fragments cloned into vectors and stored individually in host cells, and is a valuable resource for molecular cloning, gene physical mapping, and genome sequencing projects. To take the best advantage of a DNA library, a good screening method is needed. After describing pooling strategies and issues that should be considered in DNA library screening, here we report an efficient colony multiplex quantitative PCR-based 3-step, 3-dimension, and binary-code (3S3DBC) method we used to screen genes from a planarian genomic DNA fosmid library. This method requires only 3 rounds of PCR reactions and only around 6 hours to distinguish one or more desired clones from a large DNA library. According to the particular situations in different research labs, this method can be further modified and simplified to suit their requirements.
ISSN:1932-6203
DOI:10.1371/journal.pone.0116997