RNA isolation from isolatedislets and RT-PCR
Posted 15 July 2001 - 09:00 PM
Posted 28 December 2004 - 03:15 AM
As I understand you are performing RT with specific primers which you have proved effective for brain tissue. Are you sure that they are suitable for the RNA isolated from pancreatic islets?
Secondly, did you checked your isolated RNA? You can easily do it by denaturing electrophoresis.
Third, why not adding RNase inhibitor since the beginning, in the RT buffer.
Last, in case of using either oligo dT or P6 primers, be careful of their concentration vs your RNA concentration. If too concentrated, P6 will direct revers-transcription of too short products that preclude further annealing of specific PCR primers.
Because your description is limited, if still unsuccessful, please come back with more details to allow us to figure out what was wrong with your reaction. Every step is important.
Posted 06 July 2012 - 02:49 AM
I'm also having some problems with extraction of RNA from isolated islets...
Posted 06 July 2012 - 03:13 PM