ISOLATION OF DICTYOSTELIUM-DISCOIDEUM CYTOKINESIS MUTANTS BY RESTRICTION ENZYME-MEDIATED INTEGRATION OF THE BLASTICIDIN-S RESISTANCE MARKER
ISOLATION OF DICTYOSTELIUM-DISCOIDEUM CYTOKINESIS MUTANTS BY RESTRICTION ENZYME-MEDIATED INTEGRATION OF THE BLASTICIDIN-S RESISTANCE MARKER
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DOI：
10.1006/bbrc.1994.2880
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发表时间：
1994-12-30
影响因子：
3.1
通讯作者：
SUTOH, K
中科院分区：
文献类型：
--
作者：
ADACHI, H;HASEBE, T;SUTOH, K
我们已经开发了一种改进的REMI（限制酶介导的整合）系统，用于快速有效地产生突变的网骨藻细胞，用于系统筛选胞质分裂突变体。通过该系统，从2，000个网柄藻转化体中分离出三个生长为巨大多核细胞的胞质分裂突变体。对这些突变体的Southern印迹分析显示，单个拷贝的标签DNA被整合到每个基因组中。在两端具有侧翼基因组DNA的标签从突变体中的一个中被拯救并重新引入亲本Ax2菌株中。被拯救的基因和网柄藻基因组的同源重组导致了胞质分裂突变体预期的表型变化。(C)1994年出版社出版。
We have developed an improved REMI (restriction enzyme-mediated integration) system for generating mutant Dictyostelium cells quickly and efficiently for systematic screening of cytokinesis mutants. By means of this system, three cytokinesis mutants that grow as giant and multinucleate cells were isolated from 2,000 Dictyostelium transformants. Southern blot analysis of these mutants revealed that a single copy of the tag DNA was integrated into each genome. The tag with flanking genomic DNA at both ends was rescued from one of the mutants and reintroduced into the parental Ax2 strain. Homologous recombination of the rescued gene and the Dictyostelium genome led to the phenotypical changes expected for cytokinesis mutants. (C) 1994 Academic Press, Inc.