Kaufmann Y, Milcarek C, Berissi H, Penman S
Proc Natl Acad Sci U S A. 1977 Nov;74(11):4801-5. doi: 10.1073/pnas.74.11.4801.
Poly(A)+ and poly(A)- mRNA from HeLa cells were separated and translated in heterologous messenger-dependent protein synthesizing systems. Two-dimensional electrophoretic analysis revealed three classes of polypeptides. At the level of detectability in the electropherograms, a small number (about 10) of proteins were detected only among the poly(A)- mRNA products, a larger number (about 40) were produced by both poly(A)- and poly(A)+ mRNA, and a large number of polypeptides were found exclusively in the poly(A)+ mRNA products. The major product of both poly(A)+ and poly(A)- mRNA was shown to be the beta form of actin.Previous cross hybridization measurements suggested little homology between poly(A)+ and poly(A)- mRNA populations. In view of the apparent identity of many poly(A)- products with those of poly(A)+, the homology between poly(A)+ and poly(A)- mRNA sequences was examined in greater detail. cDNA complementary to only the most abundant poly(A)+ message sequences was prepared. About 10% of this cDNA hybridized to abundant sequences in the poly(A)- fraction. This corresponded to only 2% of the total mass of poly(A)+ mRNA and accounted for the failure to detect cross hybridization in previous experiments. Thus, a small number of poly(A)+ sequences appear to be present in relatively high concentration in poly(A)- mRNA as evidenced by both the translation products and the cross hybridization results.
从HeLa细胞中分离出多聚腺苷酸(Poly(A))阳性和阴性的mRNA,并在异源信使依赖性蛋白质合成系统中进行翻译。二维电泳分析揭示了三类多肽。在电泳图谱的可检测水平上,仅在Poly(A)阴性mRNA产物中检测到少量(约10种)蛋白质,Poly(A)阴性和阳性mRNA均产生较多数量(约40种)的蛋白质,并且大量多肽仅在Poly(A)阳性mRNA产物中发现。Poly(A)阳性和阴性mRNA的主要产物均显示为肌动蛋白的β形式。先前的交叉杂交测量表明,Poly(A)阳性和阴性mRNA群体之间几乎没有同源性。鉴于许多Poly(A)阴性产物与Poly(A)阳性产物明显相同,因此对Poly(A)阳性和阴性mRNA序列之间的同源性进行了更详细的研究。制备了仅与最丰富的Poly(A)阳性信使序列互补的cDNA。该cDNA的约10%与Poly(A)阴性部分中的丰富序列杂交。这仅相当于Poly(A)阳性mRNA总质量的2%,并解释了先前实验中未能检测到交叉杂交的原因。因此,如翻译产物和交叉杂交结果所示,少量Poly(A)阳性序列似乎以相对高的浓度存在于Poly(A)阴性mRNA中。