Method for 96-well M13 DNA template preparations for large-scale sequencing.
Efficient preparation of DNA templates is an important step in large-scale DNA sequencing. The 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.
Andersson
B
B
Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX 77030, USA. bjorna@bcm.tmc.edu
Lu
J
J
Edwards
K E
KE
Muzny
D M
DM
Gibbs
R A
RA
D017104:Bacteriophage M13 / Q000235:genetics*; D004279:DNA, Viral / Q000737:chemistry; D005822:Genetic Vectors / Q000096:biosynthesis / Q000302:isolation & purification; D011092:Polyethylene Glycols / Q000378:metabolism; D016133:Polymerase Chain Reaction; D017422:Sequence Analysis, DNA / Q000379:methods*; D013698:Templates, Genetic
D004279:DNA, Viral; D011092:Polyethylene Glycols