The identification of rare diseases is increasingly possible through genome sequencing, a method that involves breaking down human DNA into smaller segments known as short reads. Christian Gilissen, Lisenka Vissers, and their team discovered that a newer technique utilizing long reads is even more adept at uncovering intricate causes. Their findings indicate that eighty to ninety percent of cases can be detected, as mentioned in the American Journal of Human Genetics.
Rare diseases often stem from genetic origins. These genetic roots are progressively being traced using genome sequencing methodologies. Genome sequencing surveys an individual’s entire genome, which comprises more than 3 billion DNA building blocks.
Missed Information
When sequencing a genome, the DNA is initially divided into smaller fragments of about 300 nucleotides, referred to as short reads. These fragments are analyzed and then pieced together to recreate the complete genome. Christian Gilissen, a Professor of Genome Bioinformatics at Radboudumc notes, “This approach has enabled us to find the cause of many rare diseases.” However, the short reads technique comes with limitations. The vast array of tiny genetic fragments cannot always be accurately reassembled, particularly in extensive repetitive sequences of DNA (repeats) and segments where there have been deletions, insertions, or rearrangements (translocations). To address these challenges with short reads, various supplementary methods are often used to identify potential gaps in the data.
Long Reads
In recent years, a novel sequencing approach utilizing long reads has gained traction, particularly through advancements like PacBio HiFi sequencing. This method enables the analysis of genetic segments up to 20,000 nucleotides long, compared to the shorter segments employed in traditional reads. Vissers, a Professor of Translational Genomics at Radboudumc explains, “Initially, this technique lacked accuracy and was quite costly, but it has now become trustworthy and economically viable. This led us to question: could we substitute short reads and their accompanying tests with long reads? Would long reads be sufficient to identify enough genetic variations to replace the current methodologies in clinical settings?”
Over Ninety Percent
Gilissen and his team examined 100 samples where the genetic origins of rare diseases had been previously challenging to uncover with short reads and additional testing. In this instance, they solely relied on long reads for analysis. Gilissen mentions, “Using long reads, we were able to immediately identify 83 percent of the causes. After further investigation, we discovered that an extra 10 percent were also detected by the technique but weren’t automatically recognized. For the remaining 7 percent, we might need additional advancements in our approach. Overall, we are now capturing 93 percent of the causes that are typically difficult to identify with existing short read methods. For less complex cases, this success rate is likely even greater.”
Switching Genes On and Off
Long reads also excel at directly mapping methylation, a process that regulates whether genes are activated or deactivated through chemical alterations to the DNA. Gilissen remarks, “Short reads can’t accomplish this, necessitating a separate test. We demonstrated this with one of our samples. Ordinarily, the mother’s gene is active while the father’s gene is not, but in this case, the patient inherited no gene from the father and two from the mother, both of which were inactive. This condition was the cause of this rare disease. It’s somewhat intricate, but we successfully identified this unusual and complicated instance on the spot.”
One Test
In recent years, extensive research has been directed toward the clinical usage of long reads. Vissers notes that this was the first instance to compare 100 known, challenging-to-diagnose rare disease cases with long reads directly. The results are encouraging, she states. “With advancements in technology and decreasing costs, long reads seem poised to become the preferred approach for diagnosing rare disorders in terms of quality, user-friendliness — requiring just one test instead of several — and cost-effectiveness.”
