title: Magnetic, microplate-format plasmid isolation protocol for high-yield, sequencing-grade DNA.
abstract: We have developed a rapid, microplate-format plasmid isolation procedure to purify sequencing-grade DNA templates for high-throughput DNA sequencing operations. A modified lysozyme/boiling method is used to produce a plasmid-containing supernatant that is then purified by iron bead capture. After binding, the beads are pelleted in a magnetic field, washed and the DNA eluted in water. The method yields up to 10 micrograms plasmid DNA from a 1-mL overnight culture in a deep-well microplate. The procedure is suitable for large-scale experiments, amenable to automation and does not require expensive reagents or equipment. The entire protocol can be completed in as little as 2 h, and one technician with a 96-well pipetting station can process up to 48 plates per day. This protocol is ideal for any high-throughput operation in which template quantity, quality and reproducibility are of primary importance.
authors.affiliation: Lawrence Livermore National Laboratories, CA, USA. evans@bioworks.com
authors.forename: E W
authors.initials: EW
authors.lastname: Skowronski
authors.forename: N
authors.initials: N
authors.lastname: Armstrong
authors.forename: G
authors.initials: G
authors.lastname: Andersen
authors.forename: M
authors.initials: M
authors.lastname: Macht
authors.forename: P M
authors.initials: PM
authors.lastname: McCready
mesh_terms: D004247:DNA / Q000302:isolation & purification*; D010957:Plasmids; D017422:Sequence Analysis, DNA
chemical_list: D004247:DNA