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采用核酸酶保护测定 (NPA)和定量反转录竞争性多聚酶链反应方法 (RT- PCR) ,测定了表达重组促卵泡刺激素受体 (FSHR)的转染细胞 (Y1)的 FSHR m RNA的稳定性和半衰期。对照组 FSHR m RNA量在 8h内保持恒定水平 [NPA,(2 .9± 0 .3) pg/ μg RNA;RT- PCR,(2 .7± 0 .3) pg/ μg RNA]。用 [3H]尿苷结合到 RNA的方法 ,观察到 Actinomycin D(Act D,5 μg/ ml)在 1h内可抑制 m RNA合成达 90 %。在 Act D存在的条件下 ,FSHR m RNA量在 1h后出现一个时间依赖性的显著下降 ,用 NPA测得的半衰期为 (3.6± 0 .2 ) h,RT- PCR为 (3.1± 0 .1) h。提示m RNA的降解速率在 FSHR基因稳定表达维持上起着非常重要的作用。
FSHR m RNA of transfected cells (Y1) expressing recombinant human follicle stimulating hormone receptor (FSHR) was assayed by nuclease protection assay (NPA) and quantitative reverse transcription competitive polymerase chain reaction (RT-PCR) Stability and half-life. The amount of FSHR m RNA in the control group remained constant within 8 h [NPA (2.9 ± 0.3 pg / μg RNA; RT-PCR, (2.7 ± 0.3) pg / μg RNA]. Using [3H] uridine binding to RNA, it was observed that Actinomycin D (Act D, 5 μg / ml) inhibited the synthesis of m RNA by 90% within 1 h. In the presence of Act D, the amount of FSHR m RNA showed a significant time-dependent decrease after 1 h. The half-life measured by NPA was (3.6 ± 0.2) h, and the RT-PCR was (3.1 ± 0.1 h. It suggested that the degradation rate of m RNA plays a very important role in the stable expression of FSHR gene.