MEASURING EXPRESSION: DNA microarrays (above, left) consist of nucleic acid probes affixed to a surface. First, RNA is extracted from samples and converted into complimentary DNA (cDNA), which is labeled with fluorescent tags (1). Next, labeled cDNA fragments hybridize with the nucleic acids on the array (2). Scanning the microarray measures the fluorescence level at each spot, revealing levels of gene expression (3). In RNA-seq, RNA is also extracted from samples, fragmented, and converted into cDNA in preparation for sequencing (A). Next, the cDNA library is sequenced (B). The resulting reads are mapped to the genome and gene expression is quantified (C). © EVAN OTO/SCIENCE SOURCE
For 20 years, DNA microarrays have been the go-to method for revealing patterns of gene expression. But as the price of next-generation sequencing continues to fall, RNA sequencing (RNA-seq) has become an increasingly popular method for assessing transcriptomes.
A DNA microarray consists of a predetermined assortment of nucleic acid probes attached to a surface. To assess gene expression, researchers derive complementary DNA (cDNA) from cellular RNA, label the cDNA with a fluorescent marker, wash labeled cDNA over the array, and use lasers to assess how much cDNA has stuck to each probe. RNA-seq relies on converting RNA into a cDNA library and then directly sequencing the cDNA.
While learning how to deal with the raw data RNA-seq produces can be tricky, RNA-seq has capabilities that ...
