High-throughput plasmid purification for capillary sequencing.
The need for expeditious and inexpensive methods for high-throughput DNA sequencing has been highlighted by the accelerated pace of genome DNA sequencing over the past year. At the Joint Genome Institute, the throughput in terms of high-quality bases per day has increased over 20-fold during the past 18 mo, reaching an average of 18.3 million bases per day. To support this unprecedented scaleup, we developed an inexpensive automated method for the isolation and purification of double-stranded plasmid DNA clones for sequencing that is tailored to meet the more stringent needs of the newer capillary electrophoresis DNA sequencing machines. The protocol is based on the magnetic bead method of solid phase reversible immobilization that has been automated by using a CRS-based robotic system. The method described here has enabled us to meet our increases in production while reducing labor and materials costs significantly.
Elkin
C J
CJ
Department of Energy Joint Genome Institute, Walnut Creek, California 94598, USA.
Richardson
P M
PM
Fourcade
H M
HM
Hammon
N M
NM
Pollard
M J
MJ
Predki
P F
PF
Glavina
T
T
Hawkins
T L
TL
D001483:Base Sequence; D004270:DNA, Circular / Q000235:genetics / Q000302:isolation & purification; D019075:Electrophoresis, Capillary / Q000295:instrumentation / Q000379:methods*; D005822:Genetic Vectors / Q000737:chemistry / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D010957:Plasmids / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D017422:Sequence Analysis, DNA / Q000295:instrumentation / Q000379:methods*
D004270:DNA, Circular