GENE REPLACEMENT IN DICTYOSTELIUM - GENERATION OF MYOSIN NULL MUTANTS
GENE REPLACEMENT IN DICTYOSTELIUM - GENERATION OF MYOSIN NULL MUTANTS
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DOI：
10.1002/j.1460-2075.1989.tb03453.x
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发表时间：
1989-03-01
期刊：
影响因子：
11.4
通讯作者：
SPUDICH, JA
中科院分区：
文献类型：
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作者：
MANSTEIN, DJ;TITUS, MA;SPUDICH, JA
真核黏菌盘基网柄霉（Dictyosteelium discoideum）具有单个常规肌球蛋白重链基因（mhcA）。通过同源重组消除mhcA基因。构建了两个基因置换质粒，每个质粒携带G418抗性基因作为选择性标记，并且侧翼为0.7 kb的5“编码序列和0.9 kb的3”编码序列，或者1.5 kb的5“侧翼序列和1.1 kb的3”侧翼序列。用这两种质粒中的任一种转化后获得肌球蛋白无效突变体（mhcA-mhc）。mhcA-细胞在遗传学上是稳定的，能够进行各种运动过程.我们的研究结果提供了遗传学证据，在网骨藻的常规肌球蛋白基因所需的悬浮生长，正常的细胞分裂和孢子形成，并说明如何基因靶向可以作为一种工具，在网骨藻。
The eukaryotic slime mold Dictyostelium discoideum has a single conventional myosin heavy chain gene (mhcA). The elimination of the mhcA gene was achieved by homologous recombination. Two gene replacement plasmids were constructed, each carrying the G418 resistance gene as a selective marker and flanked by either 0.7 kb of 5'' coding sequence and 0.9 kb of 3'' coding sequence or 1.5 kb of 5'' flanking sequence and 1.1 kb of 3'' flanking sequence. Myosin null mutants (mhcA- cels) were obtained after transformation with either of these plasmids. The mhcA- cells are genetically stable and are capable of a variety of motile processes. Our results provide genetic proof that in Dictyostelium the conventional myosin gene is required for growth in suspension, normal cell division and sporogenesis, and illustrate how gene targeting can be used as a tool in Dictyostelium.