calcium phosphate transfection questions - (Apr/18/2008 )
Hi,
Really appreciate it if anyone can help me with the following questions (I'm new to this area):
1. Will you be able to see cloudiness of solution containing DNA and calcium phosphate precipitate with the naked eye? If you can't, does it mean that precipitation had failed? How to ensure if precipitation is successful?
2. Most said that you'll be able to see a layer of white covering (DNA-CaPO4 precipitate) on cells after transfection. Is it obvious under light microscope?
3. Will cloudiness be seen if DNA is absent from calcium phosphate precipitate? Is there minimal amount of DNA required for higher transfection efficiency?
Thanks.
will be continued....
-BioWizard v0.0.1-
QUOTE (BioWizard v0.0.1 @ Apr 18 2008, 03:33 AM)
Hi,
Really appreciate it if anyone can help me with the following questions (I'm new to this area):
1. Will you be able to see cloudiness of solution containing DNA and calcium phosphate precipitate with the naked eye? If you can't, does it mean that precipitation had failed? How to ensure if precipitation is successful?
2. Most said that you'll be able to see a layer of white covering (DNA-CaPO4 precipitate) on cells after transfection. Is it obvious under light microscope?
3. Will cloudiness be seen if DNA is absent from calcium phosphate precipitate? Is there minimal amount of DNA required for higher transfection efficiency?
Thanks.
will be continued....
Really appreciate it if anyone can help me with the following questions (I'm new to this area):
1. Will you be able to see cloudiness of solution containing DNA and calcium phosphate precipitate with the naked eye? If you can't, does it mean that precipitation had failed? How to ensure if precipitation is successful?
2. Most said that you'll be able to see a layer of white covering (DNA-CaPO4 precipitate) on cells after transfection. Is it obvious under light microscope?
3. Will cloudiness be seen if DNA is absent from calcium phosphate precipitate? Is there minimal amount of DNA required for higher transfection efficiency?
Thanks.
will be continued....
Usually I create my mix in a 14 mL polypropylene tube. If you turn the tube at an angle and place it over the light of a microscope, you should be able to see a cloudy column where the light hits it. This is usually visible within 5 mins of adding the HeBS2X to your calcium chloride and buffered water. I'm not sure if you will still see the cloudiness if DNA is absent. I have a feeling you probably can, but don't quote me.
-helcatt77-