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Efficient Sequencing Method Devised to ID Cells, Biomarkers

Brain visualization image

(Univ of California, San Francisco)

6 August 2014. Researchers at University of California in San Francisco and Fluidigm Corp. in South San Francisco developed a more efficient process for analyzing genetic material, still revealing cell types and biomarkers that previously required more in-depth analysis. The team led by Arnold Kriegstein, director of UCSF’s regeneration medicine and stem cell center, and postdoctoral researcher Alex Pollen, published its findings this week in the journal Nature Biotechnology (paid subscription required).

The UCSF/Fluidigm team employed a device made by Fluidigm, a developer of life science analytical tools, to prepare single-cell samples for sequencing messenger RNA or mRNA — the molecules in the genome transcribing DNA into proteins that give instructions to cells for performing basic life functions. Sequencing reveals the structure of the mRNA molecules that make it possible to identify the properties of cells performing these functions.

Current sequencing methods require deep analysis of samples, up to 5 million reads of individual cells, which makes the process costly, time-consuming, and needing larger specimen samples to analyze. The researchers, using Fluidigm’s single-cell prep system for mRNA analysis, were able to reduce the degree of analysis by 2 orders of magnitude, from 5 million to 50,000 reads per cell.

The team used microfluidics — lab-on-a-chip devices — to capture single cells, then analyzed mRNA transcriptions of 301 cells from 11 separate populations in the brain with the shallower sequencing methods.  The sequencing process, which required larger numbers of cells to study, revealed more diverse types including early developmental cells, such as radial glial cells that are progenitors for neurons and glia in the brain.

“In addition to exploring the consequences of low-depth analysis,” says Pollen in a Fluidigm statement, “the paper includes a pilot study in the brain that reveals some new biology about nervous system development.”  The researchers found in the analysis two types of early growth proteins, EGR1 and FOS, that serve as targets for a basic signaling pathway in human cell development, which had not been previously identified in the radial glia of mice.

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