2. What is your experience in performing microRNA research?
We have been studying sex differences in transcript profiles using microarray and then we are curious about the potential sex differences in the small RNA fraction. Therefore we started a screening using miRCURY LNA™ microRNA Array, and then we validated our results using miRCURY LNA™ microRNA PCR system.
3. For what type of research were you using detection of microRNA by real-time PCR? And for what part have you been using the miRCURY LNAmicroRNA PCR system?
We chose miRCURY LNA™ real-time PCR for the validation for microarray results and also to explore the potential regulation of the selected miRNAs. Northern blot was not considered since it is less accurate for quantification.
Since we performed the microarray experiment using miRCURY LNA™ microRNA Array, it seems natural to validate the results using miRCURY LNA™ microRNA PCR system.
4. What is your experience in performing quantitative PCR experiments before? We have been studying the expression levels of many genes using SYBR® Green coupled with primers designed by ourselves. I have also performed miRNA quantification using TaqMan microRNA assays.
5. What do find are the main benefits of the miRCURY LNA™ microRNA PCR system?
The miRCURY LNA™ microRNA PCR system is easy to use and provides high sensitivity and reproducibility. It makes my validation faster, easier and more quantitative. With a small amount of miRNA, I could confirm my microarray results and explore the regulation by different treatments. I personally like the reaction mix protocol since the volumes to pipet provide good margin for potential pipetting error. It makes it easier to work on a large number of samples. The running protocol is fast, less than two hours including dissociation curve. The high sensitivity gives me the control to dilute my samples to adjust the optimal amount and I find the risk of obtaining false negative results being much smaller than with other systems.
6. Did you validate your studies by other means?
We are using real-time PCR kits to validate our microarray data. There are some plans to further validate it using in situ hybridization.
7. When will we hear more about your studies?
I hope you can hear more from our study shortly. We are in the process of submitting a paper with all our findings in a peer-reviewed journal.