Glass bead purification of plasmid template DNA for high throughput sequencing of mammalian genomes
- 1 April 2002
- journal article
- Published by Oxford University Press (OUP) in Nucleic Acids Research
- Vol. 30 (7), 32e-32
- https://doi.org/10.1093/nar/30.7.e32
Abstract
To meet the new challenge of generating the draft sequences of mammalian genomes, we describe the development of a novel high throughput 96-well method for the purification of plasmid DNA template using size-fractionated, acid-washed glass beads. Unlike most previously described approaches, the current method has been designed and optimized to facilitate the direct binding of alcohol-precipitated plasmid DNA to glass beads from alkaline lysed bacterial cells containing the insoluble cellular aggregate material. Eliminating the tedious step of separating the cleared lysate significantly simplifies the method and improves throughput and reliability. During a 4 month period of 96-capillary DNA sequencing of the Rattus norvegicus genome at the Baylor College of Medicine Human Genome Sequencing Center, the average success rate and read length derived from >1 800 000 plasmid DNA templates prepared by the direct lysis/glass bead method were 82.2% and 516 bases, respectively. The cost of this direct lysis/glass bead method in September 2001 was approximately 10 cents per clone, which is a significant cost saving in high throughput genomic sequencing efforts.This publication has 18 references indexed in Scilit:
- High-Throughput Plasmid Purification for Capillary SequencingGenome Research, 2001
- Rat Genome Spurs an Unusual PartnershipScience, 2001
- Simple and rapid preparation of plasmid template by a filtration method using microtiter filter platesNucleic Acids Research, 1997
- A “Double Adaptor” Method for Improved Shotgun Library ConstructionAnalytical Biochemistry, 1996
- A High-Throughput Plasmid DNA Preparation MethodAnalytical Biochemistry, 1994
- A procedure for the large-scale isolation of highly purified plasmid DNA using alkaline extraction and binding to glass powderAnalytical Biochemistry, 1982
- A rapid boiling method for the preparation of bacterial plasmidsAnalytical Biochemistry, 1981
- Recovery of DNA segments from agarose gelsAnalytical Biochemistry, 1980
- Preparative and analytical purification of DNA from agarose.Proceedings of the National Academy of Sciences of the United States of America, 1979
- [10] Elution of DNA from agarose gels after electrophoresisMethods in Enzymology, 1979