Some genetic sequencing misses out large parts of the genome

By Dr. Ananya Mandal, MDJan 7 2020

A new study looked at the patient samples that have been earlier genetically sequenced and found that more than a quarter of genes are missed in these procedures.

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This could mean the genetic disorder detection could be flawed, say the researchers. The results of the new study titled, "Clinical Exome Studies Have Inconsistent Coverage," have been published in the latest issue of the journal Clinical Chemistry.

Researchers from the UT Southwestern Medical Center have explained that children undergoing complete genetic sequencing may be missing out on a huge chunk of their DNA from being analyzed completely.

Missed segments of patient DNA samples

They looked at patient samples from three laboratories in the United States and found that whole exome sequencing or whole genome sequencing of the patients misses out on large segments of DNA on a routine basis. This gap may prevent adequate detection of several genetic ailments, including cancers and epilepsy, they write.

The results from the samples of three labs were reanalyzed, and it was found that the labs had examined 34, 66, and 69 percent of the DNA only in their sequencing. The team explained that there had been reports of differences in the testing standards from different labs, and this has not been documented before.

Jason Park, one of the co-authors of the study and associate professor of pathology at UT Southwestern, said, "Many of the physicians who order these tests don't know this is happening. Many of their patients are young kids with neurological disorders, and they want to get the most complete diagnostic test. But they don't realize whole exome sequencing may miss something that a more targeted genetic test would find."

The results

The results showed:

• Lab A – Used VCRome v2.0 (Roche Sequencing Solutions) or xGen Exome Research Panel v1.0 (Integrated DNA Technologies) and covered 69 percent of the genes (12,184 of 17,723 genes)
• Lab B – Used VCRome v2.1 (Roche) and covered 66 percent of the genes (11,687 of 17,723 genes)
• Lab C – Used SureSelect XT2 All Exon v4 (Agilent) or Clinical Research Exome (Agilent) and covered 34 percent of the genes (5,989 of 17,723 genes)

Park explained that for whole genome sequencing or exome sequencing, the labs routinely look at the protein-producing genes of the samples, and from these, they tend to pinpoint the genetic mutations that could lead to disease.

He added that there are around 18000 exomes that need to be sequenced, and thus many can be overlooked or missed out. He warned that nearly half of the tests fail to locate a mutation. This new study looked at why many of these tests come back with negative results, he said.

Why are parts of the genome being overlooked?

For this study, they looked at patient samples for 36 patients between 2012 and 2016. Of these samples, 12 had been analyzed primarily by three national clinical laboratories, and their results had been widely varied. The authors of the study found that the genome of the patients was not fully analyzed because the labs were not following a uniform industry-advised and accepted protocol for the analysis.

Adequate analysis of the exomes includes sequencing each segment that codes for a protein at least twenty times per test. Among all the 36 samples, less than 1.5 percent of the genes were completely analyzed, and one lab particularly revealed that 28 percent of the genes were left unexamined. This lab examined only 5 percent of the DNA that was always analyzed. Yet another lab worked consistently on 27 percent of the genome and missed out on the rest, the team wrote.

What could this mean for diagnosing disease?

Park said, "And things really start to fall apart when you start thinking about using these tests to rule out a disease. A negative exome result is meaningless when so many of the genes are not thoroughly analyzed."

Jason Park, UT Southwestern

The team explained that when trying to detect a genetic condition such as epilepsy, they noted that one of the labs examined three-quarters of the whole genome in one patient sample while it examined only 40 percent of the sample in three other samples. Another lab examined less than 20 percent of the whole sequence, they wrote.

The team wrote, "The use of average measurements in large datasets such as exome/genome is only meaningful when the average is further described in combination with other summary statistics such as standard deviation and coefficient of variation... Additional statistics such as genes completely covered will also allow a more accurate assessment of the test's validity and would pressure clinical laboratories to achieve higher consistency."

What needs to be done to improve this?

Garrett Gotway, corresponding and first author of the study and a clinical geneticist at UT Southwestern, said, "When we saw this data we made it a regular practice to ask the labs about coverage of specific genes. I don't think you can expect complete coverage of 18,000 genes every time, but it's fair to expect 90 percent or more."

He added that this new study would help physicians become more aware and ask labs to look at the genome more thoroughly before pronouncing negative results.

Clinical exomes can be helpful in complex cases, but you probably don't need one if a kid has epilepsy and doesn't have other complicating clinical problems. There's a decent chance the exome test will come back negative and the parents are still left wondering about the genetic basis for their child's disease."

Garrett Gotway, UT Southwestern

The way out, he suggested, would be a complete analysis of a small panel of genes associated with the disease. This would save on costs and also raise the possibility of a diagnosis, he said.

"Although genomic testing is complex, traditional quality-control tools such as control charting may be useful in visually demonstrating the importance of intersample sequencing assessment for entire genes or specific variant locations," the authors wrote.

They concluded, "Poor consistency in complete gene coverage was seen in the clinical exome laboratories surveyed. The degree of consistency varied widely between the laboratories."