International Mammalian Genome Society

The 14th International Mouse Genome Conference (2000)

D17. High-Throughput Purification Method of Dye Terminators-Labeled Products of DNA Sequencing Reaction by Using 384 Well PCR Plates

Katsunori Aizawa1, Kazuhiro Shibata1, Masami Muramatsu1 and Yoshihide Hayashizaki1,2
1Genome Exploration Research Group, RIKEN Genomic Science Center (GSC) and Genome Science Laboratory, RIKEN Tsukuba Institute, Core Research of Evolutional Science and Technology (CREST), Japan Science and Technology Corporation (JST), Tsukuba, Ibaraki, 305-0074, Japan,
Medical School, Tsukuba University, Tsukuba, Ibaraki 305-8575, Japan.

RISA sequencer was developed by obtaining the cooperation study between CREST (JST), Shimadzu Co., and this laboratory. In this system, 384 samples could be analyzed simultaneously. We also developed previously RIKEN GS384 thermalcycler, which can perform the temperature-cycling procedures for sequencing chemistries in 4 sets of 384 well PCR plates. However, no cheap and high-throughput method has been developed to eliminate free dyes in dye terminators-labeled products. Ethanol/sodium acetate/EDTA precipitation procedure was optimized for efficient eliminating free dyes in 384 well PCR plates after dye terminators-labeling (BigDye terminator or DYEnamic ET terminator). From the initiation of the terminators-labeling with RIKEN GS384 to the end of the purification steps, the same PCR plate can be used without transferring each sample to another plate. The method would be also suited for automation on liquid-handling workstations.

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