Meghaan M. Ferreira Ph.D. Contributor GEN and Clinical OMICs

Breaking Through Barriers between Benchtop and Bedside

The discovery of PCR in 1983 revolutionized molecular biology and changed how clinicians diagnose infectious diseases and genetic disorders. Today, new advances in PCR technology, including more sensitive assays and high-throughput instruments, may further revolutionize healthcare by enabling routine blood tests for early cancer detection and changing how doctors prescribe medications based on an individual’s genetics.

“It would be wonderful if, when you go for your annual checkup and the doctor takes a blood sample to check your cholesterol, they could also look for the presence of DNA fragments from cancer cells,” imagined Fred Kramer, Ph.D., Professor of Microbiology, Biochemistry & Molecular Genetics, Rutgers New Jersey Medical School. Dr. Kramer’s laboratory focuses on developing extremely sensitive and selective PCR assays: “we want them to be quantitative, we want them to be multiplexed, but above all we want them to be simple and inexpensive.”

The laboratory’s most recent endeavor has focused on enhancing the sensitivity of PCR assays to enable selective amplification of cellfree circulating tumor DNA (ctDNA) present in a blood sample to detect somatic cancer-related mutations. Dr. Kramer’s lab developed what they call “SuperSelective” primers to amplify these extremely rare DNA fragments, and they have successfully used them to detect as few as ten mutant fragments among 1,000,000 wild-type sequences—even when they only differed by a single nucleotide polymorphism (SNP).

SuperSelective primers have a unique three-part design that enables this selectivity: a 20–24 nucleotide-long “anchor” sequence that binds both wild-type and mutant fragments on the 5’ end, a short seven-nucleotide-long “foot” sequence that perfectly complements the mutant sequence without binding the wild-type on the 3’ end, and a “bridge” sequence designed not to bind any regions in the target DNA between the two ends.

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