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Generation of mismatch repair-profi cient cell lines from human HCT116 cells
https://repo.qst.go.jp/records/69993
https://repo.qst.go.jp/records/69993b0d55f99-c5be-419f-bd0f-31e4d37c798d
| Item type | 会議発表用資料 / Presentation(1) | |||||
|---|---|---|---|---|---|---|
| 公開日 | 2009-12-14 | |||||
| タイトル | ||||||
| タイトル | Generation of mismatch repair-profi cient cell lines from human HCT116 cells | |||||
| 言語 | ||||||
| 言語 | eng | |||||
| 資源タイプ | ||||||
| 資源タイプ識別子 | http://purl.org/coar/resource_type/c_c94f | |||||
| 資源タイプ | conference object | |||||
| アクセス権 | ||||||
| アクセス権 | metadata only access | |||||
| アクセス権URI | http://purl.org/coar/access_right/c_14cb | |||||
| 著者 |
Shiomi, Naoko
× Shiomi, Naoko× Mori, Masahiko× Imai, Takashi× Shiomi, Tadahiro× 塩見 尚子× 森 雅彦× 今井 高志× 塩見 忠博 |
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| 抄録 | ||||||
| 内容記述タイプ | Abstract | |||||
| 内容記述 | We have used HCT116 as a parental cell line for targeted disruption of DNA repair related genes, because HCT116 has many advantages for gene-disruption studies (such as near dipoloid karyotype, stable chromosome number). However, HCT116 has a disadvantage that is the defect in mismatch repair (MMR) due to a point mutation (from serine [TCA] to stop [TAA] at amino acid residue 252) in both allele of one of MMR genes, hMLH1. In order to rescue the MMR-defect in HCT116 cells, we tried to replace the mutated portion of hMLH1 gene with normal hMLH1 fragment using the gene-replacement vector constructed with the normal hMLH1 fragment. The vector also has neo gene as a dominant selective marker. After introducing the vector DNA into HCT116 cells by electroporation, we selected G418-resistant clones. Out of these clones we selected the clones in which mutated portion of hMLH1 were replaced with normal sequence examining by PCR and Southern blotting. Furthermore normal size of hMLH1 protein was detected in these clones. Because MMR-deficient cells have been reported to show 10 times more resistance to the drug, 6-thioguanine (6TG), and also show more than 100 times higher spontaneous mutation frequency, we examined the hMLH1-proficient cells for the sensitivity to 6TG and spontaneous mutation frequencies at HGPRT locus. These cells showed normal sensitivity to 6TG and normal levels of spontaneous mutation frequency indicating that these cells were normal in MMR function. |
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| 会議概要（会議名, 開催地, 会期, 主催者等） | ||||||
| 内容記述タイプ | Other | |||||
| 内容記述 | The 32nd Annual Meeting of the Molecular Biology Society of Japan | |||||
| 発表年月日 | ||||||
| 日付 | 2009-12-12 | |||||
| 日付タイプ | Issued | |||||
