Notice & News
The Standard Sensitivity RNA Analysis Kit (15 nt) is used for the automated assessment of the quality of high concentration RNA samples. The efficient analysis of RNA is crucial to many applications, including next-generation sequencing technologies; qPCR, microarray analysis, and Northern blot analysis. To further enhance the ease of sample analysis post-electrophoresis, PROSize® 2.0 calculates the RNA Quality Number (RQN), providing a quantitative sample quality metric for downstream applications. With a quantitative measure of RNA quality and high-throughput capability, this kit is ideal for most RNA analysis applications.
The efficient and accurate analysis of RNA samples is enabled by the use of a high quality RNA ladder. In the electropherogram below, a typical separation of the Standard Sensitivity RNA Ladder is depicted. The reproducibility of the Standard Sensitivity RNA Ladder ensures the quality analysis of total RNA samples.
Figure 1. A typical separation of the Standard Sensitivity RNA Ladder (DNF-382) performed on a Fragment Analyzer equipped with a Short Capillary Array (33-55) using the Standard Sensitivity RNA Analyzer Kit (DNF-471)
As shown in Figure 2, a total RNA separation, as viewed via an electropherogram, has three elements: the small RNA region, the small rRNA subunit peak, and the large rRNA subunit peak. For ease of viewing, the large rRNA subunit peak is highlighted purple in PROSize®, while the small rRNA subunit peak is highlighted pink. The size of the small and large rRNA subunits varies at all levels of biological hierarchy, from Domain to Species. As such, the sizes of the small and large rRNA subunits shown in the RNA Property Summary are the most common sizes for prokaryotic, eukaryotic, and plant rRNA. The RQN provides a quantitative measure of total RNA quality. An RQN of >7 indicates the sample is ideal for downstream applications including NGS.
Figure 2. Two separations of high quality total RNA extracted from yeast. Teh separation was performed using the Standard Sensitivity RNA Analysis Kit on a Fragment Aanlyzer equipped with a Short Capillary Array (33-55). Each separation has three elements used in the calculation of RQN; the small RNA region, the small rRNA subunit, and the large rRNA subunit.
Poor quality total RNA is readily identified by this kit as shown in Figure 3. The top panel depicts a total RNA sample with an RQN of 6.2. Sequencing of a sample such as this could result in decreased read counts and poor quality sequencing results overall. The bottom panel depicts a completely degraded total RNA sample not suitable for sequencing. As RNA quality decreases, the quality of sequencing results declines.
Figure 3. Two separations of low quality total RNA. The separations were performed using the Standard Sensivity RNA Analysis Kit (15nt) on a Fragment Analyzer equipped with a Short Capillary Array (33-55). The top panel shows a total RNA Sample from rice root that falls below the minimum RQN quality threshold of 7. The bottom panel shows a completely degraded total RNA sample from yeast.
Accurate quantification is important to the assessment of sample quality. The Standard Sensitivity RNA Analysis Kit (DNF-471) provides equivalent quantification of RNA compared to common methods of fluorometric quantification. A one to one linear relationship is observed (Figure 4) between concentrations calculated by a fluorometric instrument and the Fragment Analyzer for the same samples.
Figure 4. Concentrations measured by a fluorometric instrument were plotted against the concentrations measured by the Fragment Analyzer for the same total RNA Sample. The observed one to one linear relationship supports the equivalency of quantification between the two instruments.