笔记详情
标题
Real-Time RT-PCR
内容
Real-Time RT-PCR
Total RNA was extracted from cells using RNeasy kit (Qiagen) and was quantified by UV absorbance spectrophotometry. The reverse transcription reaction was performed by using the Superscript First-Strand Synthesis System (Invitrogen) in a final volume of 20 μl containing 2 μg of total RNA, 100 ng of random hexamers, 1× reverse transcription buffer, 2.5 mM MgCl2, 1 mM dNTP, 10 U of RNaseOUT, 20 U of Superscript reverse transcriptase, and DEPC-treated water. Quantitative real-time RT-PCR analyses of human KEAP1, NRF2, GCLc, GCLm, GSR, PRDX1, GSTA3, GSTA2, NQO1, MRP1, and MRP2 were performed by using Assay-on-Demand primers and probe sets from Applied Biosystems. Assays were performed by using the ABI 7000 Taqman system (Applied Biosystems). β-actin was used for normalization.
点击翻译
来源
PLoS Med. 2006 October; 3(10): e420.
类别
关键词