transfection

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valind
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transfection

Post by valind » Wed Oct 08, 2008 1:43 am

hi,everyone
I am new in the research work.how can I kown the efficiency of transfection when I transfect plasmid or miRNA in the cells?
thanks!

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biohazard
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Post by biohazard » Wed Oct 08, 2008 1:13 pm

In general you should include some sort of reporter gene on your construct, and then see the outcome when you cultivate your microbe - e.g. the colour of your plaque or the ability of a colony to utilise a nutrient the target organism normally cannot use. That should allow you to calculate the relative efficiency of the transfection, as well as to identify & collect the transfected organisms. Most good biotechnology textbooks and many Internet resources contain classical examples of these. That is, if we're talkng about, say, bacterial, viral or yeast expression systems.

You can of course transfect immortalised B-cells and whatever, but these are a bit different story, though the principle is the same: you need something in your construct that allows you to identify and select the desired cells. Again, this can be something that can be readily visualised (like GFP) or only permits transfected cells to survive (an ability to metabolise certain nutrients or toxins that the wild type cells cannot, for example).

And of course you need to verify that the cells produce whatever the construct is originally supposed to produce, and not only express your reporter gene!

sam2008
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Re: transfection

Post by sam2008 » Thu Oct 16, 2008 6:56 am

The original meaning of transfection was 'infection by transformation', i.e. introduction of DNA (or RNA) from an eukaryote virus or bacteriophage into cells, resulting in an infection. Because the term transformation had another sense in animal cell biology (a genetic change allowing long-term propagation in culture, or acquisition of properties typical of cancer cells), the term transfection acquired, for animal cells, its present meaning of a change in cell properties caused by introduction of DNA.
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