denaturing gel for RNA - (Dec/05/2008 )
Hi all...
Planing to do Northern hybridisation so standardising with Denaturing gel or formaldehyde gel for RNA from shrimp tissue.
Iam uploading a picture of the gel i did. It is said to get 28s and 18s bands but i have a single band and smear to the bottom of the gel. i ran the gel for 3hrs. Can anybody help me out to find wats the problem? please do tell if there is DNA contamination on seeing the gel.
Thankyou
can somebody tell me how to upload the gel picture...dont find any keys...
Reply to your original post. At the bottom of the text box you type into, you will see "Attachments". In small letters underneath "Manage Current Attachments" it says "select a file" in small print. To the right of this, it says "Choose file". Click this, choose your file, and click the green upload button. The maximum is 3 MB
Ulterior motive: I want to see what advice people give you, and what your gel looks like
What is your preparation protocol for your gel? Qiagen bench guides (RNA section) provides a very good guide on preparing and troubleshooting denaturing formaldehyde gels. don' t run your gel too long.
did you denature your RNA before loading? did you soak the gel in the buffer at least 30min to equilibrate before loading?
Hi all....
i have uploaded the gel picture. I did denature the RNA before loading and ran the gel in buffer without addition of the sample for 30min so as to equilibriate before loading. i followed the protocol according to Sambrook or Maniatis Denaturing formaldehyde gel. well something amiss in the gel i feel. is it fine as iam seeing only single band when there shud be 28s and 18s as it is eukaryotic....
Thank you
Hi all....
i havent got reply to the query...anyone can help out.
Thanku
i havent got reply to the query...anyone can help out.
Thanku
Look like that's genomic DNA and degraded RNA. How long was the run? For checking RNA you could simply run a non-denaturing gel at 120V for 30-40min.
Hi ...
I ran the gel for 3hrs....but iam onto doing Northern hybridisation so juz wanted to know whether the gel picture i have uploaded is apt but since there is only one band instead of the two 18s and 28s...iam having doubts whether wat i did is rite???if there is any suggestions they are mostly welcome....
thankyou