Van Ness J, Hahn W E
Nucleic Acids Res. 1980 Sep 25;8(18):4259-70. doi: 10.1093/nar/8.18.4259.
Mouse liver poly(A)+mRNA was reverse transcribed using oligo-p(dT) or random oligonucleotides as primers to yield cDNA about equal to the mass of the template RNA. The size profile of the oligo-p(dT)-primedd cDNA was similar to that of the template RNA. RNA or cDNA driven saturation annealing of labeled single copy genomic DNA (scDNA) showed that 2% of the scDNA was complementary in either case indicating the sequence complexity of cDNA was equivalent to that of the template mRNA. These results establish for the first time that cDNA represents essentially all of the sequence complexity of a diverse template RNA population in which individual mRNA species are present in vastly different concentrations. RNA driven hydridization of the cDNA showed that about 40% of the cDNA mass represents most of the sequence complexity of the template RNA. Also, kinetics of this hybridization indicate a complexity of 58,000 kb for the template RNA, a value similar to that obtained by scDNA hybridization. We conclude that appropriately characterized cDNA probes can be used to make valid qualitative and quantitative comparisons of the complex, infrequent class mRNAs of different cells and tissues.
使用寡聚 - p(dT) 或随机寡核苷酸作为引物对小鼠肝脏多聚腺苷酸加尾（poly(A)+）mRNA 进行逆转录，以产生质量约等于模板 RNA 的 cDNA。以寡聚 - p(dT) 为引物合成的 cDNA 的大小分布与模板 RNA 相似。RNA 或 cDNA 驱动的标记单拷贝基因组 DNA（scDNA）饱和退火表明，在这两种情况下，2% 的 scDNA 是互补的，这表明 cDNA 的序列复杂性与模板 mRNA 相当。这些结果首次证实，cDNA 基本上代表了不同模板 RNA 群体的所有序列复杂性，其中各个 mRNA 种类的浓度差异极大。cDNA 的 RNA 驱动杂交表明，约 40% 的 cDNA 质量代表了模板 RNA 的大部分序列复杂性。此外，这种杂交的动力学表明模板 RNA 的复杂性为 58,000 kb，该值与通过 scDNA 杂交获得的值相似。我们得出结论，经过适当表征的 cDNA 探针可用于对不同细胞和组织中复杂且罕见的 mRNA 类别进行有效的定性和定量比较。
