Microinjecting cells using a constant-flow microinjection system.
Microinjecting cells using a constant-flow microinjection system.
复制标题
使用恒流显微注射系统显微注射细胞。
DOI：
10.1101/pdb.prot5590
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发表时间：
2011
影响因子：
--
通讯作者：
Gasiorowski,JoshuaZ
中科院分区：
文献类型：
--
作者：
Dean,DavidA;Gasiorowski,JoshuaZ
有效地将外源DNA导入细胞进行表达和功能研究是现代细胞生物学的一项基本技术，而通过显微注射直接输送遗传物质仍然是一种可靠的转基因方法。在这里描述的系统中，从移液管的尖端输送恒定流量的样品，并且注入到池中的样本量取决于移液管在池中停留的时间。该系统可以使用市售组件以相对较低的成本进行组装，这些组件通常包括可调节两种压力(背压和注射压力)的压力调节器、毛细管座以及粗糙和精细的微操作器。恒流系统在设计上比脉冲流系统注入的可再生细胞体积要少。然而，在实践中，这种差异可以忽略不计。
INTRODUCTIONThe efficient delivery of exogenous DNA to cells for expression and function studies is an essential technique of modern cell biology, and direct delivery of genetic material by microinjection remains a reliable means of transfection. In the system described here a constant flow of sample is delivered from the tip of the pipette, and the amount of sample injected into the cell is determined by how long the pipette remains in the cell. The system can be assembled at relatively low cost using commercially available components: Typically, a pressure regulator that can be adjusted for two pressures (back pressure and injection pressure), a capillary holder, and coarse and fine micromanipulators. A constant-flow system, by design, injects less reproducible cell volumes than does a pulsed-flow system. However, with practice, this difference can be made negligible.