Genomics Application Notes

and benign cells micro-dissected from FFPE tissue blocks. Differential expression of the target miR 1 46a, and the reference RNU48, was observed across the samples.

RNA yields were between 4 1 ng to 72 ng (Figure 1 ). All samples show similar fragmented RNA quality from FFPE samples (Figure 2).

Fig. 1. Summary of RNA yield from 3 pairs of tumor/benign samples.

Fig. 3. miR146a and RNU48 miRNA gene expression.

Conclusions The data from this study shows that the Beckman Coulter FormaPure Kit can be used for low-input sample extraction such as LCM from FFPE tissue blocks. The magnetic bead-based extraction protocol provides automation-friendly, scalable throughput and does not require vortexing, centrifugation, or filtration steps. Acknowledgement The data used in this application note was provided by a customer.

Fig. 2. Typical fragmented RNA from micro-dissected FFPE samples.

MicroRNA Gene Expression Indicating that miRNA was Successfully Extracted from LCM Samples 1 0 ng of total RNA was used for miR 1 46a and RNU48 gene expression to determine miRNA gene expression. The average cycle threshold (Ct) was calculated from triplicates of each tumor/benign pair. The average Ct value for miR 1 46a target gene expression in benign cells was 32.22±0. 11 and 3 1 .67±0.22 in tumor cells. The average Ct value for RNU48 miRNA reference gene expression was 24. 1 7±0. 1 6 in benign cells and 24. 1 4±0.0 1 3 in tumor cells (Figure 3). This result indicates that downstream RT-PCR of target and reference miRNAs is feasible with robust Ct values derived from RNA, extracted utilizing the FormaPure method from tumor

Author Bee Na Lee, PhD, Staff Application Scientist Beckman Coulter Life Sciences

The FormaPure reagents are not intended or validated for use in the diagnosis of disease or other conditions. © 2014 Beckman Coulter, Inc. All rights reserved. Beckman Coulter, the stylized logo, SPRI, Agencourt, and FormaPure are registered trademarks of Beckman Coulter, Inc. and are registered with the USPTO. All other trademarks are the property of their respective owners.

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