Case Study Demonstrates Importance of MIQE for qPCR Data Analysis
News Feb 17, 2012
The study examined the effects of RNA sample quality and reference gene stability — two critical components of the Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) checklist — on gene expression, comparing differences between normal and tumor breast cancer samples.
“These data demonstrate the importance of the MIQE methodology on the final conclusions drawn from qPCR experiments,” said Sean Taylor, lead author of the case study and a Bio-Rad senior field application specialist. “The guidelines were designed to offer a standardized, methodical approach to this highly sensitive technique to ensure that all data published with qPCR are consistent, reproducible, and — most important — a true reflection of the conclusions.”
As a result of its high sensitivity, qPCR has become the gold standard for validating DNA microarray data and is routinely used to determine gene expression differences in a wide variety of samples. However, inadequate sample processing and handling, poor primer design and validation, and inappropriate reference gene selection may result in data being misinterpreted. The MIQE guidelines were published to assist the scientific community to produce consistent, high-quality data from qPCR experiments.
The case study (Bio-Rad Tech Note 6245) used the minichromosome maintenance (MCM) protein MCM7 as a model target gene to investigate the importance of appropriate reference gene selection and RNA sample quality from breast cancer samples as described by MIQE. Following the MIQE guidelines, a significant increase in gene expression of MCM7 was observed between normal and tumor samples when using high-quality and -purity RNA with normalization using stable reference genes. However, inconclusive and even opposite results were obtained when using poor-quality RNA samples and unstable reference genes. These results demonstrate that inappropriate conclusions may be drawn from qPCR data if key steps from the MIQE guidelines are ignored.
Mechanism Controlling Multiple Sclerosis Risk IdentifiedNews
Researchers at Karolinska Institutet have now discovered a new mechanism of a major risk gene for multiple sclerosis (MS) that triggers disease through so-called epigenetic regulation. They also found a protective genetic variant that reduces the risk for MS through the same mechanism.
Antarctic Worm and Machine Learning Help Identify Cerebral Palsy EarlierNews
A research team has released a study in the peer-reviewed journal BMC Bioinformatics showing that DNA methylation patterns in circulating blood cells can be used to help identify spastic cerebral palsy (CP) patients. The technique which makes use of machine learning, data science and even analysis of Antarctic worms, raises hopes for earlier targeted CP therapies.
Ancient Syphilis Genomes Decoded for First TimeNews
Researchers recovered three genomes of the bacterium Treponema pallidum from skeletal remains from colonial-era Mexico, and were able to distinguish the subspecies that causes syphilis from the subspecies that causes yaws. It was not previously thought possible to recover DNA from this bacterium from ancient samples.