24-hour age difference causes twice as much gene expression divergence as 100 generations of adaptation to a novel environment

Gene expression profiling is one of the most reliable high-throughput phenotyping methods, allowing to quantify the transcript abundance of expressed genes. Because many biotic and abiotic factors influence gene expression, it is recommended to control them as tightly as possible. Here we show that the 24-hour age difference of Drosophila simulans females that were subjected to RNA-Seq five and six days after eclosure results in more than 2000 differentially expressed genes. This is twice the number of genes that changed gene expression during 100 generations of evolution in a novel hot laboratory environment. Importantly, most of the genes differing in expression due to age introduce false positives/negatives if an adaptive gene expression analysis is not controlled for age. Our results indicate that tightly controlled experimental conditions, including precise developmental staging, are needed for reliable gene expression analyses, in particular in an evolutionary framework.