3次元マイクロアレイシステムによる
K-rasコドン12の変異解析に関する基礎的検討
前 川 真 人*1 谷 口 照 美*2 立 林 千 夏*3 堀 井 俊 伸*4
竹 下 明 裕*5 椙 村 春 彦*6 菅 野 康 吉*7 米 川 裕 之*8
長 岡 智 紀*9 菅 野 剛 史*10
Basic Studies on Mutation Analysis of K-ras Codon 12
by Use of Three-Dimensional Microarray System
Masato MAEKAWA, MD*1, Terumi TANIGUCHI*2, Chika TATEBAYASHI*3,
Toshinobu HORII, MD*4, Akihiro TAKESHITA, MD*5, Haruhiko SUGIMURA, MD*6,
Kokichi SUGANO, MD*7, Hiroyuki YONEKAWA*8,
Tomonori NAGAOKA*9 and Takashi KANNO, MD*10
A next-generation DNA microarray system, FD10 has been developed. It is
based around the PamChip, a custom-made microarray, which consists of a
solid three-dimensional structure that facilitated the incorporation of
probe molecules. We applied this microarray system on a detection of K-ras
mutation at codon 12 in some cancer cell lines. The PCR products amplified
by use of FITC labeled primers were applied onto probe-absorbed microarray.
After hybridization, the signal was imaged by CCD camera and analyzed by
the exclusive software. We confirmed the microarray results by PCR-SSCP
and sequencing analyses.
Ten, two and three out of 15 cell lines were homozygous for wild type allele,
heterozygous for wild and mutant allele, and homozygous for mutant alleles,
respectively. Signals hybridized with antisense probes were stronger than
those with sense probes, without PSN1 cell line. The system had a good
reproducibility. Essentially, the microarray results were consistent with
PCR-SSCP and sequencing results.
In conclusion, the FD10 microarray system was easy to operate and short to get results. It might be useful for a focused array applicable for specific purposes. The K-ras mutation detection system worked well and will be applied to clinical specimens soon.
[Rinsho Byori 51 : 306～312, 2003]
*1Department of Laboratory Medicine, Hamamatsu University School of Medicine, Hamamatsu 431-3192
【Key Words】microarray(マイクロアレイ)，chip(チップ)，three dimension(三次元)，K-ras mutation(K-ras 変異)
受付2003年1月15日・受理2003年3月19日
*1～5浜松医科大学医学部臨床検査医学，*6同 病理学第一，*10同 副学長(〒431-3192
浜松市半田山1-20-1)
*7栃木県立がんセンター研究所 がん遺伝子研究室・がん予防研究室(〒320-0834
宇都宮市陽南4-9-13)
*8,9オリンパス光学工業㈱ゲノム医療事業推進室(〒192-8512 八王子市久保山町2-3)
E-mail :mmaekawa@hama-med.ac.jp