Specimens

Pools (>10 animals) of mixed sexes rat pineal glands or retinas were prepared from tissue obtained at six time points during a 24-hour period. Animals were housed in a 14:10 light:dark lighting cycle. Samples were collected at mid-day (ZT7) and mid-night (ZT19), and at one hour before and one hour after the light-to-dark transition (ZT13 and ZT15) and the dark-to-light transition (ZT23 and ZT1). In addition, a mixed tissue sample was prepared from material collected from three rats at ZT7 and at ZT19. The tissues were: cortex, cerebellum, midbrain, hypothalamus, hindbrain, spinal cord, retina, pituitary, heart, liver, lung, kidney, skeletal muscle, small intestine, adrenal gland.

RNA Preparation

Samples of tissue from the three animals were mixed. Total RNA was extracted using an RNeasy Kit with on-column DNase treatment. To prepare the mixed tissue sample, equal amounts of RNA from each of the 15 tissues were combined.

RNA-Seq libary preparation and sequencing

Total RNA (10 ug) was polyA-selected, fragmented and barcoded. Samples were run six per sequencing lane on an Illumina HiSeq2000 machine, generating paired-end 101-mer reads.

Note: In some cases, the mixed tissue values for selected genes are exceptionally high because that gene is very highly expressed in only one tissue, e.g., Rho and GH are strongly expressed only in the retina and pituitary gland, respectively.

Bioinformatics methods

See bioinformatics methods for details.