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Researchers Put Turtles in Their Place with a MicroRNA-Based Approach

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A new technique for animal classification could quell the long-running debate between paleontologists and molecular biologists as to where turtles belong on the evolutionary tree. While some camps have argued that turtles are closer to crocodilians and birds, researchers at the Mount Desert Island Biological Laboratory in Bar Harbor, Maine; Dartmouth College; and Harvard University have published data in Biology Letters suggesting that turtles are more closely related to tuataras and lizards. The team identified 77 new microRNA families in the Anolis carolinensis lizard genome, four of which are also expressed in Chrysemys picta bellii, the Western painted turtle, leading them to conclude that turtles and lizards have much more in common than previously thought.

"When we first started studying microRNAs, we noticed that they seemed to cluster animals hierarchically, and thus they could be used as a new source of phylogenetic information," says Kevin Peterson, an associate professor at Dartmouth. "We've spent several years developing the proof of concept, and have published several studies where microRNAs were used to help elucidate phylogenetic inter-relationships, including jawless fishes, arthropods, annelids, brachiopods, and sponges."

Peterson says the real challenge was sorting through all of the data, as the lizard genome was much larger than others they had previously analyzed. "These deep sequencing approaches give, ultimately, millions of small RNA reads and we needed a way to wade through all these data to find known and novel microRNAs," he says. "This was especially true working with vertebrate small RNA libraries, as we map the sequencing reads to a genome assembly and fold an interval of sequence that encompasses the mapped position."

Peterson credits MDIBL bioinformatician Ben King with significantly aiding the effort by tweaking the software used to analyze the miRNA data. King developed the program miRMiner, which helped the team find known and novel miRNAs in small RNA libraries.
The group is currently working on applying this miRNA analysis approach to several other projects, including a study of the inter-relationships among placental mammals, lizards, and ecdysozoan phyla — a group of protostome animals that was first segregated using 18S ribosomal RNA genes in 1997.

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