TaqMan reagents speed up workflows and enable analysis of trace amounts of RNA in cell and tumour samples
Research related to a class of non-coding small RNA molecules called microRNAs (miRNAs) is one of the fastest-growing areas in biomedical research.
These molecules have been found to regulate gene expression and have also been implicated in a number of different diseases, including cancer.
As researchers continue to investigate how the activity of miRNAs impacts the formation and progression of cancer, they will need more advanced tools.
Responding to that need, Applied Biosystems has developed a new set of sophisticated miRNA analysis tools designed to determine the activity, or profile the expression levels, of large numbers of miRNA molecules from trace amounts of RNA sample.
These new TaqMan-based tools require the smallest amount of starting genetic material needed for miRNA expression profiling applications compared to existing miRNA analysis tools.
Use of these new tools is expected to streamline miRNA analysis workflows for researchers and provide comprehensive coverage of miRNAs expressed in cancer cell line and tumour samples, advancing scientists' understanding of the different roles that miRNAs play in the development and progression of cancer.
Previous research has shown that by profiling expression patterns of miRNAs, scientists can identify potential miRNA biomarkers that classify different disease types and identify disease susceptibility.
In addition, miRNA expression profiling is helping researchers to understand the nature of cancer stem cells, which play a key role in the resistance of cancers to conventional therapies.
Expression profiles of miRNAs have also revealed clues about how tumours metastasise and spread to other regions of the body.
However, researchers performing these kinds of miRNA analysis applications for cancer research often face the challenge of only being able to obtain tiny amounts of RNA from cancerous samples, the starting genetic material needed to carry out these investigative studies.
These tools are currently available in limited release through an early access programme.
When used together, these tools will provide researchers with a TaqMan-based solution designed to streamline the workflow for miRNA analysis and provide broad, up-to-date coverage of known miRNAs expressed in biological samples, including samples that contain minute amounts of RNA.
"Together, these tools give researchers a powerful technology that allows them to create comprehensive profiles of miRNAs expressed in cancer samples," said Chen.
"The use of these new TaqMan miRNA analysis tools should help researchers develop biomarkers for cancer, better understand cancer stem cells, and further elucidate the role of miRNAs in the development and progression of cancer".
Today, these tools are being used by researchers enrolled in an early access programme to reveal unique miRNA expression signatures from as little as 1ng of total RNA sample.
This represents at least 100-fold less genetic material than is currently required by existing technologies.
By early summer, the Megaplex RT Pools, Megaplex PreAmp Primers and updated TaqMan MicroRNA Arrays are expected to be generally available for use in miRNA expression analysis applications of human, mouse and rat.
Cancer samples are often collected from heterogeneous tissue, with only a tiny portion in the sample being the cancerous cells containing the RNA needed for expression profiling studies.
This RNA must be obtained through either needle biopsy or a process known as laser capture microdissection (LCM).
Archived samples are often formalin-fixed paraffin embedded (FFPE), a process in which the genetic material is often degraded and RNA yield is relatively low.
Commonly, these samples are extremely valuable and of a finite nature, so being able to work with limited amounts of RNA is of paramount importance to researchers.
Use of the TaqMan Megaplex reagents is expected to enable researchers to generate a complete profile of expressed miRNA molecules from RNA obtained by needle biopsy, LCM, and freshly frozen or FFPE tumour samples.
For example, Thomas Schmittgen, an associate professor at the Ohio State University's College of Pharmacy, USA, has been studying the role of miRNAs in cancer development, with a focus on developing new cancer therapies and diagnostics based on research related to analysis of patterns of miRNA expression.
He has previously used the TaqMan MicroRNA Assays to detect the mature, biologically active form of miRNA in a study of cancer cell lines that evaluated the differences in levels of pre-cursor forms and mature forms of miRNA present in normal and cancerous cells.
As part of Applied Biosystems' early access programme, Schmittgen has been using the TaqMan Megaplex workflow in his research aimed at determining the role of miRNAs in the formation of cancer.
He has been developing miRNA expression patterns into biomarkers for potential use in diagnostic assays to detect cancer from a small amount of cells or tissue.
"We are very interested in assaying the mature miRNA expression in samples with very low cell numbers," said Schmittgen.
"The Megaplex reagents and TaqMan MicroRNA Assays allow us to accurately determine the miRNA expression profiles of these samples from very little starting material".
Another researcher who expects to advance his research from use of the TaqMan Megaplex reagents is Aaron Fletcher, a resident research fellow at the University of Iowa Hospitals and Clinics, USA.
He has been investigating how small groups of self-renewing stem cells found in cancerous tumours contribute to both recurrence and metastasis of cancer in patients.
Fletcher is using the TaqMan MicroRNA Arrays for expression profiling experiments designed to distinguish tumour cells from cancer stem cells, based on signature miRNA expression patterns found in each type of cell.
However, because of the paucity of stem cells in most cancer samples, he plans to use the Megaplex reagents to profile miRNAs in a distinct set of cancer stem cells.
"Profiling the miRNA expression of cancer stem cells will potentially have an impact on the way that we understand and treat tumours," said Fletcher.
"The Megaplex reagents and TaqMan MicroRNA Assays will help our efforts to identify cancer stem cells by allowing us to profile as little as one cell for miRNA changes that can then be correlated with established miRNA array profiles for these cells."
