Large scale library generation for high throughput sequencing |
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Authors: | Borgström Erik Lundin Sverker Lundeberg Joakim |
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Institution: | Science for Life Laboratory, Division of Gene Technology, Royal Institute of Technology (KTH), School of Biotechnology, Solna, Sweden. |
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Abstract: | BackgroundLarge efforts have recently been made to automate the sample preparation
protocols for massively parallel sequencing in order to match the increasing
instrument throughput. Still, the size selection through agarose gel
electrophoresis separation is a labor-intensive bottleneck of these
protocols.Methodology/Principal FindingsIn this study a method for automatic library preparation and size selection
on a liquid handling robot is presented. The method utilizes selective
precipitation of certain sizes of DNA molecules on to paramagnetic beads for
cleanup and selection after standard enzymatic reactions.Conclusions/SignificanceThe method is used to generate libraries for de novo and re-sequencing on the
Illumina HiSeq 2000 instrument with a throughput of 12 samples per
instrument in approximately 4 hours. The resulting output data show quality
scores and pass filter rates comparable to manually prepared samples. The
sample size distribution can be adjusted for each application, and are
suitable for all high throughput DNA processing protocols seeking to control
size intervals. |
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