Agilent Total RNA Isolation Mini Kit Application Note

About this manual

This document describes an application study evaluating the Agilent Total RNA Isolation Mini Kit (p/n 5185-6000) for the removal of residual plasmid DNA from RNA preparations isolated from transiently transfected cells. The study compares the standard kit protocol against a competitor's silica-based kit and modified versions of both protocols that include organic extraction and in-solution DNase treatment steps.

Results from quantitative real-time PCR assays indicate that the Agilent kit, even without additional DNase treatment, removes significantly more input plasmid DNA than the silica-based method. The study concludes that the Agilent kit provides superior performance for obtaining highly purified RNA suitable for accurate gene expression analysis in research applications.

What's inside

• Abstract
• Introduction
• Materials and Methods
• Plasmid-Spiked Experiments
• Transient Transfection Experiments
• Results
• Plasmid DNA Spiking Experiments
• Transient Transfection Experiments Results

Frequently asked questions

What is the maximum amount of RNA that requires an increase in enzyme and buffer amounts for digestion?

The protocol states that for more than 60 ug of RNA, the enzyme and buffer amounts should be increased accordingly.

How long must the RNA pellet sit at room temperature after removing the supernatant?

The pellets must be allowed to sit for 5 minutes at room temperature after the supernatants are removed.

What is the incubation temperature and duration for dissolving the RNA pellet with lysis solution?

Lysis solution is added and the tubes are vortexed twice to dissolve the pellets, though no specific temperature or duration is stated for this step.