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More accurate, powerful genetic analysis tool opens new gene-regulation realms
Date:4/22/2013

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The work involved a novel enrichment method, which applied a special "chemical cross-linker" to physically join the RMT to an RNA that it is trying to methylate, said Vahid Khoddami, the study's co-author and a member of the Cairns Lab. "Our new technique takes advantage of the mechanism of the enzyme. The drug/crosslinker we used looks like cytosine, so it is incorporated in place of the cytosine in the RNA. The RMT tries to methylate this drug thinking it is a normal target cytosinebut instead becomes crosslinked to the RNA, defining the precise location of the intended methylation. As our reaction-based method requires that the enzyme both bind the RNA and commit to the act of methylation, it greatly increases our identification of true positives," said Khoddami.

"This technique gives us 200-fold enrichment, when two-fold enrichment has been considered a great result in the past," said Khoddami. "In fact, for some RNA types, the enrichment is more than 700-fold."

After the enrichment process, high-throughput gene sequencing is used to analyze the RNA samples obtained.

"Our enrichment results were fantastic by themselves, but in the sequencing process we made another important discovery," Khoddami said. "We found that after sequencing, the target cytosine in the modified RNA instead appeared as an alternative molecule, guanosine, more than 50% of the time. After sequencing, you can look for these cytosine to guanosine transversions and know you have the precise targetin a single experiment."

According to Khoddami, ten cytosine RMTs are known in humans, and only two of them have been partially characterized. "None of the other eight have been studied in the laboratory," he explained, "although some of them have been shown to have connections to cancer, infertility, and particular genetic disorders in humans.

"These diseases have been puzzling because previously we did not have the tools to analyze the RNA. Now we have be
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Contact: Linda Aagard
801-587-7639
University of Utah Health Sciences
Source:Eurekalert

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