MicroJect 1000A Robotic Micro Injection System
The microinjection technique involves the direct transfer of DNA into a cell for subsequent integration and expression. This method is the dominant approach to producing transgenic mice and has a number of advantages over other methods such as electroporation and fusion, which are less efficient in delivering DNA into cells. The process requires the use of a micropipette to pierce the membrane of the target cell or embryo and insert DNA into its cytoplasm or nucleus. This method allows for the targeting of specific regions of a genome and thus is useful for testing gene function and genetic diseases.
The process is most commonly used to introduce DNA into a developing mouse egg (zygote). Following the fertilization of a female and male pronuclei, a small amount of genetic material can be introduced by microinjection into the zygotic nucleus of the fertilized egg. This results in the random integration of a foreign gene in each cell of the resulting offspring, and can be used to generate permanent transgenic lines.
Alternatively, the technique can also be used to introduce a genetic tag into the nucleus of a cell or embryo for detection purposes. This tag can then be traced by a number of methods including PCR, Southern blot hybridization and radioisotope labeling.
However, the microinjection process is extremely labor intensive. Only a limited number of cells can be injected at a time and many injections are required for successful transgene expression. Additionally, a significant percentage of injected embryos will be irreversibly damaged and lysed following the microinjection process (see Fig. 8.8A).
To overcome these limitations, the development of a high-throughput quantitative microinjection system is essential. Currently, manual quantitative microinjection is only feasible for larger-sized cells such as mouse embryos and oocytes.21 However, for human cells that can range from 7 mm to 25 mm in size, a system that can deliver high-throughput and quantitative control is needed.
The MicroJect 1000A is designed to meet these needs. This system uses a 3-DOF robot manipulator to hold the microinjection pipette and a sterilized cell holder chip with multiple cell trapping channels. A PC with full automation programming controls the system to provide positive injection pressure and negative cell trapping pressure for each individual cell.
A unique feature of this system is the ability to hold the cell or oocyte stationary while performing the injection using a separate pressure channel. This allows the user to precisely control the injection volume while minimizing the risk of diluting samples by capillary action. Furthermore, the system has a secondary balance pressure that provides a constant, controlled injection volume irrespective of capillary forces. This feature is especially important when working with pico volumes such as those required for nuclear microinjections into oocytes or early stage embryos. This is especially critical when introducing CRISPR/CAS9 plasmids into cells to increase genome editing efficiency.micro injection