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Analytical Chemistry

Direct RNA Sequencing

Helicos BioSciences reports a method for direct RNA sequencing that avoids first converting RNA to complementary DNA

by Celia Henry Arnaud
September 28, 2009 | APPEARED IN VOLUME 87, ISSUE 39

RNA sequencing usually requires an indirect approach of converting the RNA to complementary DNA and then sequencing that DNA. This cDNA approach doesn't work well for RNA samples that are short, degraded, or available in limited quantities. Now, scientists at Helicos BioSciences, in Cambridge, Mass., report a method for direct RNA sequencing (Nature, DOI: 10.1038/nature08390). Patrice M. Milos and coworkers extend the company's single-molecule DNA-sequencing methods by incorporating a polymerase that can use RNA as its template. The target RNA molecules are arrayed on a planar surface, and their sequences are determined by synthesizing the complementary strand through repeated additions of nucleotides with fluorescent labels and cleavable inhibitors that block the addition of other nucleotides. The researchers take a picture of the array after each addition to find out where that particular nucleotide was incorporated. They subsequently remove the inhibitors and add the next nucleotide, repeating the cycle as many times as desired until the entire sample has been sequenced. Compared with hybridization-based sequencing methods, the new approach is less prone to biases and sequencing errors, the researchers note.



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