International Mammalian Genome Society

The 14th International Mouse Genome Conference (2000)

Table of Contents * Genes, Physical Mapping and Sequencing * Gene Identification * Bioinformatics * Common Resources * RIKEN Session * Functional Genomics * Complex Traits * Mutagenesis * Developmental Genetics and Differentiation * Verne Chapman Memorial Lecture I and II

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 Hayashizaki^1,2
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,
²Medical 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.