Construction of a shuttle vector for the filamentous fungus Neurospora crassa.
Construction of a shuttle vector for the filamentous fungus Neurospora crassa.
复制标题
丝状真菌粗糙脉孢菌穿梭载体的构建。
DOI：
10.1073/pnas.80.4.1058
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发表时间：
1983
影响因子：
11.1
通讯作者：
Lambowitz,AM
中科院分区：
文献类型：
--
作者：
Stohl,LL;Lambowitz,AM
我们已经构建了一个重组质粒，帕尔斯-1，在链孢菌和大肠杆菌中自主复制。帕尔斯-1由来自脉孢菌菌株P405-Labelle的线粒体质粒、脉孢菌qa-2+基因和E. coli质粒pBR 325。帕尔斯-1转化脉孢菌qa-2+基因的频率比主要通过整合转化的质粒高5- 10倍。当E.用来自脉孢菌属转化体的DNA转化大肠杆菌，我们不仅回收了帕尔斯-1，而且回收了较小的质粒帕尔斯-2，其经历了pBR 325中的大部分和可能所有的Labelle序列以及紧邻侧翼序列的缺失。帕尔斯-2似乎也在脉孢菌中自主复制，但效率低于帕尔斯-1。Southern印迹显示游离的帕尔斯-1和帕尔斯-2存在于脉孢菌转化体的核和胞质（高速离心的上清液）部分中，并且在线粒体中也可以检测到小的可变比例的质粒。帕尔斯-1和帕尔斯-2构成脉孢菌属的假定穿梭载体。
We have constructed a recombinant plasmid, pALS-1, that replicates autonomously in both Neurospora and Escherichia coli. pALS-1 consists of the mitochondrial plasmid from Neurospora strain P405-Labelle, the Neurospora qa-2+ gene, and E. coli plasmid pBR325. pALS-1 transforms the Neurospora qa-2+ gene at frequencies 5- to 10-fold higher than those for plasmids that transform mainly by integration. When E. coli was transformed with DNA from Neurospora transformants, we recovered not only pALS-1 but also a smaller plasmid, pALS-2, which had undergone deletion of most and possibly all Labelle sequences, and the immediately flanking sequences in pBR325. pALS-2 also appears to replicate autonomously in Neurospora, but less efficiently than does pALS-1. Southern blots show that free pALS-1 and pALS-2 are present in nuclear and cytosolic (supernatant from high-speed centrifugation) fractions of Neurospora transformants and that small, variable proportions of the plasmids also can be detected in mitochondria. pALS-1 and pALS-2 constitute putative shuttle vectors for Neurospora.