Dunn K, Dickstein R, Feinbaum R, Burnett B K, Peterman T K, Thoidis G, Goodman H M, Ausubel F M
Department of Genetics, Harvard Medical School, Boston 02114.
Mol Plant Microbe Interact. 1988 Feb;1(2):66-74. doi: 10.1094/mpmi-1-066.
We have cloned alfalfa nodule-specific cDNAs that code for leghemoglobin (Lb), glutamine synthetase (GS), and three unidentified nodulins. Hybrid-select translation of nodule RNA followed by 2-D gel electrophoresis showed that the Lb-specific cDNA corresponded to at least four Lb species of 12 kDa. One of the unidentified cDNA clones (N-32/34) corresponded to at least five polypeptides of 32-34 kDa; a second unidentified cDNA clone (N-14) corresponded to an individual polypeptide of 14 kDa. The in vitro translation product(s) of the RNA hybrid selected by the third unidentified cDNA clone (N-22) formed a single band at 22 kDa on a one-dimensional gel. Northern and dot blot analyses of RNA isolated from wild-type nodules and from defective nodules elicited by a variety of Rhizobium meliloti mutants showed that 1) RNAs corresponding to the Lb, nodule-specific GS, and three unidentified nodulins were coordinately expressed during the course of nodule development, and 2) all five nodulins were expressed in Fix- nodules that contained infection threads and bacteroids but were not expressed in nodules that lacked infection threads and intracellular rhizobia.
我们已经克隆了苜蓿根瘤特异性cDNA，它们编码豆血红蛋白（Lb）、谷氨酰胺合成酶（GS）以及三种未鉴定的根瘤蛋白。对根瘤RNA进行杂交选择翻译，然后进行二维凝胶电泳分析，结果表明，Lb特异性cDNA对应于至少四种分子量为12 kDa的Lb蛋白。其中一个未鉴定的cDNA克隆（N-32/34）对应于至少五种分子量为32 - 34 kDa的多肽；另一个未鉴定的cDNA克隆（N-14）对应于一种分子量为14 kDa的单一多肽。由第三个未鉴定的cDNA克隆（N-22）选择的RNA的体外翻译产物在一维凝胶上形成了一条位于22 kDa处的单带。对从野生型根瘤以及由多种苜蓿根瘤菌突变体诱导产生的缺陷型根瘤中分离得到的RNA进行Northern杂交和斑点杂交分析，结果显示：1）与Lb、根瘤特异性GS以及三种未鉴定的根瘤蛋白相对应的RNAs在根瘤发育过程中协同表达；2）所有这五种根瘤蛋白在含有感染丝和类菌体的固氮根瘤中表达，但在缺乏感染丝和细胞内根瘤菌的根瘤中不表达。
