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Competitive quantitative measurement of the AMPA receptor gene expression at the single cell level.

机译:在单细胞水平上竞争性定量测量AMPA受体基因的表达。

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Our laboratory has developed a competitive reverse transcriptase polymerase chain reaction (RT-PCR) procedure to analyse the mRNA expression of the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptor subunits in single cells. By the use of an internal RNA standard competing equally with the four subunit's mRNA, we have analysed 283 whole single hippocampus CA1 cells from adult rat brain. The cells were sampled from three groups of animals: one control group, one group subjected to preconditioning ischemia, and one group subjected to global cerebral ischemia. After reverse-transcription and PCR-amplification of mRNA in the cells, the PCR product was digested using subunit specific endonucleases and quantified by Cy-5 fluorescence. The median mRNA copy numbers achieved from control rats were 290, 247, 207, and 16 GluR1-4, respectively.
机译:我们的实验室开发了一种竞争性逆转录聚合酶链反应(RT-PCR)程序,以分析单个细胞中α-氨基-3-羟基-5-甲基异恶唑-4-丙酸酯(AMPA)受体亚基的mRNA表达。通过使用与四个亚基的mRNA均等竞争的内部RNA标准,我们已经分析了成年大鼠大脑中的283个完整的单个海马CA1细胞。从三组动物中采集细胞:一组为对照组,一组为预处理缺血,另一组为全脑缺血。在细胞中mRNA的反转录和PCR扩增后,使用亚基特异性核酸内切酶消化PCR产物,并通过Cy-5荧光定量。从对照大鼠获得的中值mRNA拷贝数分别为290、247、207和16 GluR1-4。

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