array scanning protocol
Scanning is performed with the Axon GenePix 4000B microarray scanner. GenePix pro V6.0 is used to read the raw intensity of the image.
nucleic acid hybridization to array protocol
Hy3 labeled small RNA samples hybridized at 56C for 16 hours in Exiqon miRCURY LNA Array Hybridization buffer, then washed three times in buffer A, B, C, respectively.
nucleic acid labeling protocol
Small RNAs from rat serum were labeled with Hy3 fluorescent label using Exiqon miRCURY LNA Array power labeling kit as described by the manufacturer. Briefly, small RNA samples were first treated with CIP enzyme in CIP buffer, then labeled with Hy3 in Labeling buffer with DMSO.
nucleic acid extraction protocol
Serum of rats were homogenized in RNAiso for Small RNA (Takara, Dalian, China) and the small RNA was isolated from the serum as per the manufacturers protocol. The quality of RNA was verified on NanoDrop ND1000 (Thermo Scientific, USA) and agarose gel.
Rats from two treatment groups were fed with 20uL DHA or Linoleic acid (both from Sigma-Aldrich, USA) by gentle intragastric administration every two days, and the control rats were fed with saline solution in the same way.
3-week-old female Wistar rats allowed to acclimatize to the conditions in our animal facility (constant humidity, temperature and 12 h dark/light cycle) for 1 week. Then the animals were provided with water and standard rat chow mixed with saturated fat and starch, which mimic the daily diets in modern society.