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Jo Vandesompele (left) and Jan Hellemans (right), cofounders of Biogazelle.

Commercial providers have traditionally used software to facilitate the design of qPCR assays. While these algorithms detect sequences such as single-nucleotide polymorphisms and secondary structure that may impair or bias amplification, they do not guarantee that the assay will work in the lab. To validate an assay, users must still test the resulting PCR primers for efficiency and specificity with the target sequence.

Meeting the MIQE Guidelines through Full Wet-Lab Validation
Bio-Rad collaborated with Biogazelle to design, optimize, and experimentally validate SYBR® Green-based gene expression assays for the mouse genome. PCR primer pairs for each mouse transcript were designed following strict guidelines on maximum transcript coverage, minimal overlap with known SNPs, and spanning large introns where possible.

In accordance with the MIQE guidelines, every assay was wet-lab validated following strict assay performance standards for amplification efficiency, specificity, sensitivity, and linear dynamic range. Assay specificity was confirmed using next-generation sequencing (NGS). This approach yielded validated assays for more than 90 percent of the mRNA transcriptome.

“Using Bio-Rad’s PrimePCR assays now allows users to run high-quality MIQE-compliant assays without having to spend the effort performing the lab validation themselves,” said Hellemans.

Additionally, the PrimePCR mouse assays are:

• Expertly curated — predesigned pathway and disease plates curated by Thomson Reuters for canonical pathways and related collections help researchers identify and investigate key targets
• Customizable — easy-to-use plate configurator allows users to lay out assays on a plate exactly as they choose or use a suggested plate layout template
• Fully integrated — full integration with CFX real-time PCR systems and data analysis software streamlines data collection and analysis

For more information on Bio-Rad’s PrimePCR products, please visit www.bio-rad.com/primepcr.

Consider this: Despite the limited amount of research done so far, scientists have already discovered bacteria which are capable of degrading highly toxic compounds on the seabed floor. If their power is harnessed properly, the bacteria may eventually be used as ecological biosensors or as tools to detoxify harmful substances derived from human activity.

According to Jesús María Arrieta, CSIC researcher at the Mediterranean Institute for Advanced Studies

The number of marine species used as a source of genes with commercial interests grows at a rate of 12% per annum

This fast pace of discovery will eventually open the door to multiple biotechnological applications in fields such as bioenergy, food or cosmetics.

It is in that spirit that a team of Spanish researchers, coordinated by the Spanish National Research Council (CSIC), has started to sequence the genome of the global deep ocean. The scientists havve chosen to go deeper than previous studies, penetrating waters between 200 to 4,000 meters deep in order to discover viruses, bacteria and protists that inhabit the mysterious ocean depth.

To read more on this story visit Spanish researchers sequence the genome of global deep ocean or read the original press release on the CSIC website (in Spanish).

See how this new approach is incorporated into the V3 Western Workflow™ in this short video from Bio-Rad Laboratories.

The study has revealed up to 50 replisome components, some of which had not been previously identified. The results represent the first proteomic charaterisation of the replisome opening the door for developing new therapeutic strategies to treat replication-based diseases such as cancer.

For more information read the full story on the CNIO website.

Reference:
Lopez-Contreras, Andres. (2013-04–1) A Proteomic Characterization of Factors Enriched at Nascent DNA Molecules. , 3(4), 1105-1116. DOI: 10.1016/j.celrep.2013.03.009

PCR inhibition is a frequent problem for people working with complex biological samples such as food, plant material and clinical samples. Many attempts have been made to try to overcome PCR inhibition, mainly involving some form of sample preparation. Each of these solutions however does have its limitations.

In an exclusive interview published in the European Biotechnologist in December 2010, Stephen Bustin, author of the Minimum Information for publication of Quantitative real-time PCR Experiments (MIQE) guidelines, related that

essentially every single step of the PCR assay is controversial. If I’m looking at mRNA expression, I must know something about the integrity of my RNA and if there are any inhibitors present. Now, that is not controversial. The controversial point is that people don’t actually do that: they don’t look at integrity and they don’t look for inhibition.

To minimize the impact of PCR Inhibitors, Bio-Rad Laboratories has launched the SsoAdvanced™ universal supermix which is significantly more resistant to PCR inhibitors due to the novel Sso7d fusion polymerase. This allows individual users to overcome inhibition without the usual limitations.

In a follow up interview, Dr. Bustin spoke about his use of Sso7d technology in overcoming PCR Inhibition. His reaction:

We noticed that whereas PCR reactions carried out using normal mastermixes were frequently totally inhibited, those carried out using the fast mix were considerably less so. When we carried out a 1:5 or 1:10 dilution of the crude DNA samples, virtually all samples amplified using the fast mix yielded amplification products

Read: Professor Stephen Bustin Talks About His Use of Sso7d Technology in Overcoming PCR Inhibition.

Request a sample of SsoAdvanced universal supermix today and overcome your inhibition.

The study, which appears in the journal Science, was conducted by the Social Science Genetic Association Consortium (SSGAC), which includes researchers at NYU, Erasmus University, Cornell University, Harvard University, the University of Bristol, and the University of Queensland, among other institutions.

The SSGAC conducted what is called a genome-wide association study (GWAS) to explore the link between genetic variation and educational attainment—the number of years of schooling completed by an individual and whether he or she graduated college. In a GWAS, researchers test hundreds of thousands of genetic markers for association with some characteristics such as a disease, trait or life outcome.

Because the sample included people from different countries—where markers for schooling vary significantly—the research team adopted the International Standard Classification of Education (ISCED) scale, which is a commonly used method for establishing a uniform measure of educational attainment across cohorts.

Anticipating that very large samples would be required to credibly detect genetic associations, the SSGAC researchers assembled a total sample size more than 10 times larger than any previous genetic study of any social-scientific outcome. The team examined associations between educational attainment and genetic variants called single-nucleotide polymorphisms, or SNPs, which are tiny changes at a single location in a person’s genetic code.

The study found that the genetic markers with the strongest effects on educational attainment could each only explain two one-hundredths of a percentage point (0.02 percent). To put that figure into perspective, it is known from earlier research that the SNP with the largest effect on human height accounts for about 0.40 percent of the variation.

Combining the two million examined SNPs, the SSGAC researchers were able to explain about 2 percent of the variation in educational attainment across individuals, and anticipate that this figure will rise as larger samples become available.

“We hope that our findings will eventually be useful for understanding biological processes underlying learning, memory, reading disabilities and cognitive decline in the elderly,” said co-author Daniel Benjamin, a behavioral economist at Cornell who is a co-director of the SSGAC.

“Another contribution of our study is that it will strengthen the methodological foundations of social-science genetics,” said David Cesarini, an NYU assistant professor at the Center for Experimental Social Science and the Center for Neuroeconomics, who also co-directs the SSGAC. “We used 125,000 individuals to conduct this study. Previous studies used far smaller samples, sometimes as small as 100 individuals and rarely more than 10,000. These small samples make sense under the assumption that individual genes have large effects. However, if genes have small effects, as our study shows, then sample sizes need to be very large to produce robust findings that will reliably replicate in other samples.”

The researchers were careful to note that they have not discovered “the gene for education” or that these findings somehow imply that a person’s educational attainment is determined at birth.

“For most outcomes that we study as social scientists, genetic influences are likely to operate through environmental channels that are modifiable,” explained NYU sociologist Dalton Conley, one of the study’s co-authors who also serves on the Advisory Board of the SSGAC. “We have now taken a small but important first step toward identifying the specific genetic variants that predict educational attainment. Armed with this knowledge, we can now begin to examine how other factors—including public policy, parental roles, and economic status—dampen or amplify genetic effects and ultimately devise better remedies to bolster educational outcomes.”

Thanks to New York University for this story.