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Mar 1, 2009 (Vol. 29, No. 5)

Quality Makes a Difference in RNA Prep

New Methodologies Are Being Counted On to Bring Order and Repeatability to Gene Expression

  • Oncology Focus

    “Our exclusive focus on oncology requires us to deal with a large number of samples for RNA profiling in microarrays and other large-scale platforms,” says Stephen Tirrell, Ph.D., director of molecular technologies at Millennium Pharmaceuticals, The Takeda Oncology Company. 

    “Our biggest challenge is maximizing the quality and availability of biopsy samples from the trial site,” Dr. Tirrell explains. “Getting a good biopsy from the clinic to the lab is a monumental task, demanding an all-out effort. In oncology, tumor content of the biopsy is a big variable that adds to the challenge.”

    Dr. Tirrell feels that new technologies developed for RNA isolation and purification have greatly helped to move the field forward. “The new kits are much faster and more reliable,” he states. “Before they were available we could spend a week and a half just processing the samples and isolating the material that was required to do the actual experiments.”

    A component of cancer biomarker studies is focused on a search for cancer-related biomarkers in whole blood, and much of Dr. Tirrell’s investigations are based on this approach. However blood has its limits in such investigations. “It is a good surrogate tissue for biomarker studies, however it does not directly inform you what is happening in the tumor,” he adds.

  • Amplification of Tiny Samples

    “Our technology focuses on minute amounts of RNA such as those from small cell populations, microdissection samples, fluorescence-activated cell-sorting samples, or fine-needle aspiration biopsies,” says Jeffrey Hung, Ph.D., director of marketing at SABiosciences.

    The company has introduced its RT2 Nano PreAMP cDNA Synthesis Kit, designed to take on 1 ng total RNA on PCR arrays. Using multiplex PCR-based preamplification it provides amplification of gene-specific cDNA target templates with minimal bias. With this kit one can prepare enough cDNA from each RNA sample for gene-expression analysis on as many as four different pathways, Dr. Hung says. A built-in external RNA control allows the detection of inhibitors of reverse transcription, ensuring the efficiency of the first-strand cDNA synthesis reactions.

    “The beauty of the technology is that it enables the accurate and sensitive analysis of up to four different pathway PCR Arrays (~350 genes) from as few as 100 cells or 1 ng total RNA, Dr. Hung adds.  For precious samples that can only be obtained in minute amounts, we provide an ideal solution for pathway gene-expression analysis.” 

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