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Transcriptome Subtraction Pinpoints Unknown Viruses in Human Serum

By BiotechDaily International staff writers
Posted on 02 Jul 2013
An approach based on subtracting viral DNA and RNA from that normally present in human serum and then comparing the results to databases of known viral genomes enables screening patients for the presence of unknown viruses.

Investigators at St. Louis University (MO, USA) recently described the use of the next-generation sequencing approach called transcriptome subtraction to look for unknown viral sequences in samples of serum. Transcriptome subtraction requires ultradeep sequencing to establish which DNA and RNA belong to the human genome and separate this material from extraneous viral nucleic acids.

Depth in DNA sequencing refers to the number of times a nucleotide is read during the sequencing process. Deep sequencing indicates that the coverage, or depth, of the process is many times larger than the length of the sequence under study. The term "deep" has been used for a wide range of depths (at least seven times), and the newer term "ultradeep" has appeared in the scientific literature to refer to even higher coverage (at least 100 times). Even though the sequencing accuracy for each individual nucleotide is very high, the very large number of nucleotides in the genome means that if an individual genome is only sequenced once, there will be a significant number of sequencing errors. Furthermore, rare single-nucleotide polymorphisms (SNPs) are common. Hence to distinguish between sequencing errors and true SNPs, it is necessary to increase the sequencing accuracy even further by sequencing individual genomes a large number of times.

After the human genome was identified and removed from the equation, the investigators used data from well-curated databases and advanced bioinformatic tools to eliminate DNA and RNA from all known viruses. Any nucleic acids remaining belonged to unknown viruses.

“We have discovered a technology that allows us to detect new viruses,” said contributing author Dr. Adrian Di Bisceglie, professor of internal medicine at Saint Louis University. “We isolate DNA and RNA, amplify the amount of genetic material present in the blood, do ultradeep sequencing, and use an algorithm to search for matches for every known piece of genetic code, both human and for microbes. Just as the human microbiome project is chronicling the bacteria that live and coexist in every person, we also are studying the human virome to know more about the viruses that live in all of us—we believe not all are harmful and some may even be beneficial.”

The study was published in the June 11, 2013, online edition of the journal Biochemical and Biophysical Research Communications.

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