A comparative mRNA-seq profiling of the liver, rumen villi, mammalian gland and kidney of Holstein dairy cows with high vs normal milk-urea excretion and with low vs normal (control) crude protein intake
A comparative mRNA-seq profiling of the liver, rumen villi, mammalian gland and kidney of Holstein dairy cows with high vs normal milk-urea excretion and with low vs normal (control) crude protein intake
In this study, 24 multiparous German Holstein cows were stalled in pairs for an adaptation and then 2-week experimental periods at the FBN facility. At the end of the experiment, the cows were slaughtered by a captive bolt stunning and followed by exsanguination. Tissue samples of the liver, kidney, mammalian gland and rumen villi were dissected, snap frozen in liquid nitrogen and stored until used at -80°C. Total RNA was isolated using TRI reagent (Sigma-Aldrich). Trace DNA contamination was removed by an on-column DNase I treatment (Baseline-Zero DNase), and the RNA was cleaned-up using a Nucleospin RNA kit. The RNA quantity was determined using NanoDrop and the RNA integrity was evaluated on an Agilent 2100 Bioanalyzer (Agilent Technologies).   2 µg of total RNA with RIN > 8 was used for library preparation using a TruSeq Stranded mRNA Kit with 11 cycles of PCR-amplification according to the manufacture's recommendation (Illumina).