Usually when we transfect the gene into this organism we sometimes get good expression but also sometimes bad expression. Is there any precaution or protocol to make most efficient transfection and gene expression?
It is difficult for people to offer help without details.
What is your transfection agent?
What is your delivery method?
What regions are you trying to transfect? What cell type(s)?
Is it a reporter transgene or therapeutic?
Do you want stable integration into the genome, or transient expression?
What is the mouse strain/age/gender?
Agree with the above comment. Transient methods are convenient but are subject to experimental variation, especially if you are a beginner. If the research problem is important enough, recommend generating a transgenic mouse line using a promoter that is specific to the brain cell type of interest. Literature search will give you a series of options (promoters) to choose from. In order to obtain consistent expression, one can incorporate insulators on either side of the transgene construct. The resulting mouse line will be useful for both basic as well as translational research. Of course, making a proper transgenic mouse line takes time and effort. But if the problem being addressed is important, it would be worth the effort.
Dear Dr. Jenkins My transfecting agent is lipofectamine and delivery method is intracerebral. Region is whole brain and cells are neuronal as well as non neuronal. Ihave both reporter trransgene and therapeutic both. I want stable integration. Strains are Male (8-10 weeks) wild type and CBS knockout mice.
Hello Pradip,
Have you tried using an in vivo specific delivery reagent, such as Invivofectamine 2.0? Here is a paper where researchers did siRNA delivery directly to the prefrontal cortex. http://www.jneurosci.org/content/32/13/4562.full
I'm checking with the scientist in our lab who developed the product with regards to DNA delivery, but I would think this would offer a higher efficiency of integration.
I'll reach out to you directly to discuss further. Hope this is helpful.
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