Squeezing Out More Gene-Expression Data

In the world of gene-expression profiling, microarrays reign, but they are not without drawbacks. Microarrays have trouble with low-abundance transcripts and unidentified genes, for instance. So-called open systems, such as differential display and serial analysis of gene expression, don't require prior sequence knowledge, but they also either lack sensitivity or aren't quantitative, says Sten Linnarsson of Global Genomics in Stockholm.

Global Genomics' new Tangerine whole-genome profiling system is an open technology that quantifies gene expression using competitive PCR. Total RNA is converted to cDNA and cleaved with three Type IIS restriction enzymes (which cleave outside of their recognition motifs to leave a 4-bp gene-specific overhang). Each restriction product is split into 256 subreactions, ligated to an adapter containing one of the 256 corresponding sticky ends, subjected to two rounds of PCR, and then separated by capillary electrophoresis.

Each transcript is thus potentially displayed three times (once for each restriction enzyme)...