驱动蛋白分子驱动蛋白分子Kif18A各功能域细胞定位的研究驱动蛋白分子Kif18A各功能域细胞定位的研究

目的  探讨人驱动蛋白分子Kif18A的各功能域在真核细胞中的定位。方法  构建驱动蛋白分子Kif18A野生型及各功能域（马达区、杆状区、尾区和马达区含部分杆状区）的绿色荧光蛋白表达质粒，分别转染人乳腺癌细胞MCF7，运用聚丙烯酰胺凝胶电泳和免疫印迹法验证各蛋白的表达，免疫荧光染色法分别对细胞进行微管蛋白和DNA染色，荧光显微镜下观察各绿色荧光融合蛋白的具体亚细胞定位。结果  有丝分裂间期，Kif18A蛋白分子的马达区绿色荧光沿微管分布，尾区在细胞核和微管上均有分布，杆状区则散在分布在胞质中；有丝分裂末期，Kif18A的马达区仍沿微管分布，而马达区含部分杆状区主要集中在中体部位。结论  Kif18A蛋白分子的马达区和尾区与微管共定位；尾区含核定位序列；马达区和杆状区共同作用才能使该蛋白在有丝分裂末期定位于中体。

Analyzing intracellular localization of structural domains in kinesin motor protein Kif18A

Objective To explore the intracellular localization of structural domains in kinesin motor protein Kif18A in eukaryotic cells.Methods The GFP-expressing plasmids of the Kif18A's wild type were construced and each domain including motor,stalk,tail and motor with part of stalk domain.Then these plasmids were transfected into the breast cancer cell MCF7.The GFP-fused proteins were detected by SDS-PAGE and Western blot.The immunofluorescence staining technique was used to stain microtubules and DNA in cells.The...