論文 - 詳細
| RRC ID | 10933 |
|---|---|
| 著者 | Moriya Y, Nakamura T, Okamura N, Sakaeda T, Horinouchi M, Tamura T, Aoyama N, Kasuga M, Okumura K. |
| タイトル | Comparison of synthetic DNA templates with authentic cDNA templates in terms of quantification by real-time quantitative reverse transcription polymerase chain reaction. |
| ジャーナル | Biol Pharm Bull |
| Abstract |
Synthetic DNA templates were compared with authentic cDNA templates as standards for the real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR). The single-stranded DNA template used here targeted the multidrug resistant transporter P-glycoprotein/MDR1. The double-stranded DNA template, targeting glyceraldehyde-3-phosphate dehydrogenase (GAPDH), was synthesized using an exonuclease-free large fragment E. coli DNA polymerase I. The human colon carcinoma cell line Caco-2 and human duodenum biopsies were used to prepare the authentic cDNA templates. The standard lines were comparable for the synthetic DNA templates and authentic cDNA templates. Long-term cryopreservation at -80 degrees C resulted in the destabilization of the synthetic single-stranded DNA template compared with the authentic cDNA templates in the case of MDR1, whereas for GAPDH, the stability of the synthetic double-stranded DNA template was comparable with that of the authentic cDNA templates. Even for the synthetic DNA templates, repetitive freeze-thawing resulted in destabilization, especially at lower concentrations, and degradation products might have interfered with the RT-PCR's efficiency. The synthetic DNA templates are better than the authentic cDNA templates, but more than 5 cycles of repetitive freeze-thawing should be avoided. |
| 巻・号 | 29(3) |
| ページ | 535-8 |
| 公開日 | 2006-3-1 |
| DOI | 10.1248/bpb.29.535 |
| PII | JST.JSTAGE/bpb/29.535 |
| PMID | 16508161 |
| MeSH | ATP Binding Cassette Transporter, Subfamily B, Member 1 / biosynthesis ATP Binding Cassette Transporter, Subfamily B, Member 1 / genetics ATP Binding Cassette Transporter, Subfamily B, Member 1 / metabolism Caco-2 Cells Cryopreservation DNA / chemical synthesis* DNA / chemistry* DNA, Complementary / chemistry* DNA, Neoplasm / chemistry Data Interpretation, Statistical Duodenum / chemistry Freezing Glyceraldehyde-3-Phosphate Dehydrogenases / chemistry Glyceraldehyde-3-Phosphate Dehydrogenases / metabolism Humans Male Oligonucleotides / chemical synthesis Oligonucleotides / chemistry Reproducibility of Results Reverse Transcriptase Polymerase Chain Reaction Templates, Genetic* |
| IF | 1.863 |
| 引用数 | 13 |
| WOS 分野 | PHARMACOLOGY & PHARMACY |
| オルトメトリクス指標 |
オルトメトリクス指標項目
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| 各媒体での言及数の合計 | 0 |
| リソース情報 | |
| ヒト・動物細胞 | CACO-2(RCB0988) |