Marks M D, Larkins B A
J Biol Chem. 1982 Sep 10;257(17):9976-83.
We have synthesized cDNA clones for maize zein proteins using mRNAs purified from developing endosperm. Analysis of these clones by in vitro translation of hybrid-selected mRNAs suggested differences in sequence homology among the mRNAs for the different molecular weight zein polypeptides. These differences were also apparent in restriction maps of clones corresponding to the Mr = 22,000, 19,000, and 15,000 zeins. Using radioactive cDNA inserts as probes, we measured the extent of sequence homology among zein clones with a sensitive dot hybridization procedure. By this analysis, it was possible to distinguish clones corresponding to the different molecular weight zeins at low (Tm - 49 degrees C) to moderate (Tm - 35 degrees C) criteria, while under more stringent conditions (Tm - 20 degrees C), distinctions could be made between zein sequences within a molecular weight group. This analysis distinguish three different mRNAs for each of the Mr = 22,000 and Mr = 19,000 zeins, but only one was detected for the Mr = 15,000 zein. Comparison of the nucleotide sequences of clones for the Mr = 22,000 and Mr = 19,000 zeins showed about 60% homology throughout the coding regions. This analysis also revealed the presence of short repetitive nucleotide sequences corresponding to tandem repeats of approximately 20 amino acids in both groups of proteins.
我们利用从发育中的胚乳中纯化的mRNA，合成了玉米醇溶蛋白的cDNA克隆。通过对杂交选择的mRNA进行体外翻译来分析这些克隆，结果表明不同分子量醇溶蛋白多肽的mRNA之间在序列同源性上存在差异。这些差异在对应于分子量为22,000、19,000和15,000醇溶蛋白的克隆的限制性图谱中也很明显。我们使用放射性cDNA插入片段作为探针，通过灵敏的点杂交程序测量了醇溶蛋白克隆之间的序列同源程度。通过这种分析，在低严谨度（熔解温度-49℃）到中等严谨度（熔解温度-35℃）的条件下，能够区分对应于不同分子量醇溶蛋白的克隆，而在更严格的条件下（熔解温度-20℃），可以在一个分子量组内的醇溶蛋白序列之间做出区分。这种分析在分子量为22,000和19,000的醇溶蛋白中各自区分出三种不同的mRNA，但对于分子量为15,000的醇溶蛋白只检测到一种。对分子量为22,000和19,000的醇溶蛋白的克隆的核苷酸序列进行比较，结果显示在整个编码区域中同源性约为60%。该分析还揭示了在这两组蛋白质中都存在对应于约20个氨基酸串联重复的短重复核苷酸序列。
