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New tool isolates RNA within specific cells
Date:5/18/2009

r 'listening' to host cells before and after the initiation of a disease to determine how cells respond, or, for example study healthy immune cells versus bacterially-challenged immune cells or neurons before they learn a task and after they learn a task to determine what changes in the cell are caused by the experience.

The new UO-developed tool builds on work led by co-author Michael D. Cleary, who as a doctoral student at Stanford University unveiled the T. gondii-based approach for use in analyzing RNA synthesis and decay in 2005 in Nature Biotechnology. Cleary, now a faculty member at the University of California, Merced, worked on the UO project as a postdoctoral fellowship funded by the National Institutes of Health and HHMI.

Cleary's group built its tool with the enzyme uracil phosphoribosyltransferase (UPRT), a nucleotide salvage enzyme that prepares nucleotides for incorporation into newly synthesized RNA. By altering the nucleotide analog 4-thiouracil, the UPRT enzyme caused RNA to become tagged with thiouracil (TU), allowing the "TU-tagged" RNA to be purified from untagged RNA.

In Doe's lab, Miller, Cleary and research technician Kristin J. Robinson of the UO's institutes of Neuroscience and Molecular Biology manipulated Drosophila so that they would only express UPRT in specific target cells. The group tested the new approach in embryos, larvae and adults using microarray technology to detect cell type-specific gene expression. The researchers say the method should work in other systems, including vertebrates, by using gene transfer, retroviral delivery, electrical pulses of molecules through cell membranes, or messenger RNA injection.


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Contact: Jim Barlow
jebarlow@uoregon.edu
541-346-3481
University of Oregon
Source:Eurekalert  

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