miRNA quantification - easiest/cheapest method of measuring expression of a single miRNA in differnt ce (Oct/18/2006 )
Hi Everyone
I'm new to this miRNA stuff 
and have become a little confused by the different techniques available.
I want to measure the amount of a particular miRNA in different cell lines, and possibly also after miRNA antisense / overexpression.
I have come across the AB Taqman miRNA assay and the Ambion miRVANA qRT-PCR kits. I know there is also the miRVANA detection kit but i don't fancy northerns.
I have a few questions which i hope you guys can help me with...
1. Which kit is best or are there better alternatives?
2. Should you use a control gene in qRT-PCR, and if so, which one??
Thanks for yor help...
Andrea
2. Should you use a control gene in qRT-PCR, and if so, which one??
Thanks for yor help...
Andrea
1 The best can stand the test. I haven't test any of these kits, so i can't share the experience
. 2 Actin is a common control gene in qRT-PCR wich is a house-keeping gene in animal.
http://www.protocol-online.org/forums/inde...showtopic=19999
As Rye has pointed out a previous forum covered this....
NOTE you would want to do relative quant as your microRNA may vary from cell line to cell line but to be certain that its real variation (not eg sample prep variation) you should measure relative to a miRNA or mRNA which is pretty constant in all cell lines.
use This method or this one
I'm new to this miRNA stuff
and have become a little confused by the different techniques available.I want to measure the amount of a particular miRNA in different cell lines, and possibly also after miRNA antisense / overexpression.
I have come across the AB Taqman miRNA assay and the Ambion miRVANA qRT-PCR kits. I know there is also the miRVANA detection kit but i don't fancy northerns.
I have a few questions which i hope you guys can help me with...
1. Which kit is best or are there better alternatives?
2. Should you use a control gene in qRT-PCR, and if so, which one??
Thanks for yor help...
Andrea