Konieczny A, Jensen E O, Marcker K A, Legocki A B
Mol Biol Rep. 1987;12(1):61-6. doi: 10.1007/BF00580652.
Poly(A)+ RNA isolated from root nodules of yellow lupin (Lupinus luteus, var. Ventus) has been used as a template for the construction of a cDNA library. The ds cDNA was synthesized and inserted into the Hind III site of plasmid pBR 322 using synthetic Hind III linkers. Clones containing sequences specific for nodules were selected by differential colony hybridization using 32P-labeled cDNA synthesized either from nodule poly(A)+ RNA or from poly(A)+ RNA of uninfected root as probes. Among the recombinant plasmids, the cDNA gene for leghemoglobin was identified. The protein structure derived from its nucleotide sequence was consistent with known amino acid sequence of lupin Lb II. The cloned lupin Lb cDNA hybridized to poly(A)+ RNA from nodules only, which is in accordance with the general concept, that leghemoglobin is expressed exclusively in nodules.
从黄羽扇豆（羽扇豆属，变种Ventus）根瘤中分离得到的聚腺苷酸加尾RNA（Poly(A)+ RNA）被用作构建cDNA文库的模板。双链cDNA经合成后，使用合成的Hind III连接子插入质粒pBR 322的Hind III位点。通过差异菌落杂交筛选含有根瘤特异性序列的克隆，所用探针为用从根瘤聚腺苷酸加尾RNA或未感染根的聚腺苷酸加尾RNA合成的32P标记的cDNA。在重组质粒中，鉴定出了豆血红蛋白的cDNA基因。从其核苷酸序列推导的蛋白质结构与羽扇豆Lb II已知的氨基酸序列一致。克隆的羽扇豆Lb cDNA仅与根瘤中的聚腺苷酸加尾RNA杂交，这与豆血红蛋白仅在根瘤中表达的普遍概念相符。
