Swiderski R E, Johnson S A, Larkins B A, Graham D E
Nucleic Acids Res. 1979 Aug 10;6(11):3685-701. doi: 10.1093/nar/6.11.3685.
Analyses of sequence complexities, stage specific gene expression, and mRNA sequence divergence require polysomal RNA preparations relatively free of nuclear RNA contamination. Conventional procedures for the isolation of uncontaminated polysomal RNAs which rely on sucrose density centrifugations are laborious and unsuitable for large scale isolations. We describe here a method using sequential Sepharose chromatography for isolating polysomes and polysomal RNAs depleted for nuclear RNA. Polysomes and polysomal RNAs isolated from livers of Xenopus stimulated to produce vitellogenin were capable of directing protein synthesis in vitro and showed little evidence of degradation. The polysomal RNAs contained less than 0.5% of nuclear RNA.
对序列复杂性、阶段特异性基因表达和mRNA序列差异的分析需要相对不含核RNA污染的多聚核糖体RNA制剂。依靠蔗糖密度离心分离未受污染的多聚核糖体RNA的传统方法费力且不适用于大规模分离。我们在此描述一种使用连续琼脂糖凝胶色谱法分离多聚核糖体和去除核RNA的多聚核糖体RNA的方法。从受刺激产生卵黄蛋白原的非洲爪蟾肝脏中分离出的多聚核糖体和多聚核糖体RNA能够在体外指导蛋白质合成,并且几乎没有降解的迹象。这些多聚核糖体RNA所含的核RNA不到0.5%。