Question

1. Could we quantify just the amount of mRNA in our samples by using a spectrophotometer or Nano Drop? Why?

2. Will the 260/280 ratio differ between intact RNA samples and degraded RNA samples? Why?

Ans of 2: Is it ok?? tell me ...

Yes, 260/280 ratio would differ between intact RNA samples and degraded RNA samples. For intact RNA the ratio would be 1.9-2.1. If it is degraded, the ratio would be below 1.9 or above 2.1. Degradation indicates the RNA is not pure, contaminated with either DNA or protein. If RNA contaminated with DNA, the ratio would be below 1.9. If it is contaminated with protein, the ratio would be above 2.1. So, degraded RNA samples would be different 260/280 ratio than intact RNA.

Answer 1

1) Yes, we can quantify the amount of mRNA in our samples by using a spectrophotometer or nano drop. There is no difference between RNA and DNA in absorbance measure. To quantify only RNA, samples must be DNA free, treat it with DNase followed by a Phenol/Chloroform extraction of RNA, then suspend it in buffer and then using Nanodrop or Spectrophotometer will give reliable result.

2) Your answer is correct.  Pure RNA has an A260/A280 ratio of 2.1. Value between 1.9 to 2.1 is acceptable as you too have mentioned it. The reason mentioned by you is totally correct. As contaminated by DNA will result lower ratio because of similar absorbance.value. Whereas protein contamination result opposite way.

Just add: Quantification of RNA depends on quantity, purity and integrity. Degraded RNA doesn't mean contaminated with DNA and protein, it is purity check (you have given perfect answer for purity check). For integrity of RNA sample use Bioanalyzers. Bioanalyzers use small amounts of RNA (1-2µL) and microfluidics to determine the quantity and quality of RNA samples. It determine the sizes of rRNA bands and it's integrity. If RNA is degraded then prepare a fresh pure sample of RNA for your experiment.