I have isolated RNA from PBMC of 5 different samples. After quantification , Each have different concentration. I will use it for qPCR experiments after converting it into cDNA. How much concentration RNA should be used for cDNA synthesis.
Try to use the same starting amount of material in all samples and,most importantly, check the manufacturer's suggestions for the amount needed in RNA extraction. Try not using extreme amounts(lowest or highest suggested quantities) so you dont exceed capacities of columns used in the kits. I hope you find my suggestions useful. Best of luck!
It depends upon cDNA kit you are using. Generally, 1ug conc is sufficient to make cDNA.
I agree with Fatima Irfan . It is based on cDNA synthesis kit that you use and on the expression level of the target gene. The range that you can use is 0.5ug to 1.5ug.
Best Wishes
Hi Krisna,
Take a look at your manufacturer's recommended protocol. It is recommended to use between 0.5-2ug in most cases. First check how much RNA you can use from the sample with lowest concentration and calculate volume of other samples for the same amount of RNA. You may go as low as 100ng if you don't have enough RNA between 0.5-2ug range. That should still work but you may have hard time to detect some genes that are expressed at low levels. It is always best to use the maximum amount of RNA that your cDNA kit allows.
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