How can I dilute my RNA sample for cDNA synthesis?
I'm researching for microRNA with Qiagen kit
and followings are i've done it for it so far:
- Sample: mouse granulosa cell
- RNA extrantion: mirVANA miRNA Isolation Kit
- cDNA syn: miRCURE LNA
My RNA concentration was 22ng/ul
and the protocol said to dilute template RNA sample to 5ng/ul with nuclease-free water and total amount with the water is 20ng/ul
Anyone teach me how to dilute my RNA sample to cDNA synthesis?
Hi Eunhye,
following the protocol for cDNA synthesis, you'll need to dilute the mRNA at 5ng/ul (1vol of RNA at 22ng/ul with 3.4vol of water will give you 4.4vol of RNA at 5ng/ul) and then use 4ul of your diluted sample in the reaction mixture (which is fixed at 20ul).
I hope it helps
fred
ps: 1vol = 1 volume, whatever you want.
You can add 220 uL water to the sample to obtain a 500 ng/uL solution
Always insure to dilute your RNA with either DEPC treated water or Nuclease free water.