Hi,

For your study of gene expression you do not need to equalize the amount of RNA because your are using some reference genes in you Q-PCR. For the second question I suggest to do exactly what the vendor recommend and I thing QIAGEN recommendations are good.

Good a luck

Hi,

I just want to make sure I understand. By saying equalizing the RNA concentration for cDNA synthesis,  do you mean for example, that you are using 1ug for all your runs?

If so, are you using this formula: e.g. 20ng/ul x volume= 1ug/ul, with this you must remember to convert from ng/ul to ug/ul (do so by /1000).

Also, if your RNA yield was very good, e.g. 1000ng/ul, you may want to dilute your RNA first before using it for cDNA synthesis, as this will just make it more accurate (difficult to pipette small volumes).

For the second question, I agree with Abderrahmane, QIAGEN recommendations are good. I used the RNeasy Plus from Qiagen and also tested gene expression, and it worked well. I don't know much about the QIAGEN RNase free DNase though.

Hope this helped!

Good luck!

I mean is it important that i bring all samples RNA concentration same e.g all samples concen become 500ng/ul, OR i take different RNA concentration but for cDNA synthesis take same amount e.g 1 ul?

I usually used 1ug of RNA for cDNA synthesis, and equal amounts of my cDNA also.