Open AccessMethod for 96-Well M13 DNA Template Preparations for Large-Scale Sequencing
Author(s) -
Björn Andersson,
Jing Lü,
Kimberly M. Edwards,
Donna M. Muzny,
Richard A. Gibbs
Publication year - 1996
Publication title -
biotechniques/biotechniques
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.617
H-Index - 131
eISSN - 1940-9818
pISSN - 0736-6205
DOI - 10.2144/96206st03
Subject(s) - template , dna , filtration (mathematics) , dna sequencing , primer (cosmetics) , ethanol precipitation , precipitation , bacteriophage , sequencing by ligation , chromatography , computational biology , biology , chemistry , materials science , genomic library , base sequence , nanotechnology , genetics , extraction (chemistry) , gene , escherichia coli , physics , mathematics , statistics , organic chemistry , meteorology
Efficient preparation of DNA templates is an important step in large-scale DNA sequencing. To ensure high-quality sequence data, we have prepared M13 phage DNA templates using a glass fiber-filtration method. We present the adaptation of this protocol to a 96-well format using commercially available filter plates. Two variations are described: one using polyethylene glycol precipitation and a second where the phage particles are disrupted before filtration, thus eliminating the need for precipitation. Using either of these protocols, 96 templates can be prepared in less than 2 h. Sufficient DNA for 1–2 dye primer sequencing reactions is routinely obtained from 1 mL of culture, and the resulting sequence data are of high quality.