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Adipose metabolism is an essential contributor to the efficiency of milk production, and metabolism is controlled by several mechanisms, including gene expression of critical proteins; therefore, the objective of this study was to determine how lactational state and the genetic merit of dairy cattle affects adipose tissue (AT) metabolism and mRNA expression of genes known to control metabolism. Animals of high (HGM) and low genetic merit (LGM) were fed to requirements, and weekly dry matter intake, milk production, blood glucose, and nonesterified fatty acids were measured. Subcutaneous AT biopsies were collected at -21, 7, 28 and 56 d in milk (DIM). The mRNA expression of genes coding for lipogenic enzymes [phosphoenolpyruvate carboxykinase 1 (soluble) (PCK1), fatty acid synthase (FASN), diacylglycerol O-acyltransferase 2 (DGAT2), and stearoyl-coenzyme A desaturase (SCD)], transcription regulators [peroxisome proliferator-activated receptor γ (PPARG), thyroid hormone responsive (THRSP), wingless-type MMTV integration site family, member 10B (WNT10B), sterol regulatory element binding transcription factor 1 (SREBF1), and adiponectin (ADIPOQ)], lipolytic enzymes [hormone-sensitive lipase (LIPE), patatin-like phospholipase domain containing 2 (PNPLA2), monoglyceride lipase (MGLL), adrenoceptor β-2 (ADRB2), adipose differentiation-related protein (ADFP), and α-β-hydrolase domain containing 5 (ABHD5)], and genes controlling the sensing of intracellular energy [phosphodiesterase 3A (PDE3A); PDE3B; protein kinase, AMP-activated, α-1 catalytic subunit (PRKAA1); PRKAA2; and growth hormone receptor (GHR)] was measured. Dry matter intake, blood glucose, and nonesterified fatty acid concentrations did not differ between genetic merit groups. Milk production was greater for HGM cows from 6 to 8 wk postpartum. As expected, the rates of lipogenesis decreased in early lactation, whereas stimulated lipolysis increased. At 7 DIM, lipogenesis in HGM cows increased as a function of substrate availability (0.5, 1, 2, 3, 4, or 8mM acetic acid), whereas the response in LGM cows was much less pronounced. However, the lipogenic response at 28 DIM reversed and rates were greater in tissue from LGM than HGM cows. Peak lipolytic response, regardless of DIM, was observed at the lowest dose of isoproterenol (10(-8)M), and -21 d tissue had a greater lipolysis rate than tissue at 7, 28, and 56 d. In HGM compared with LGM cows, stimulated lipolysis at 7 and 28 DIM was greater but peaked at 10(-7)M isoproterenol, suggesting differences in tissue responsiveness due to genetic merit. Regardless of genetic merit, the expression of lipogenic genes decreased markedly in early lactation, whereas those controlling lipolysis stayed similar or decreased slightly. Cows of HGM had lower expression of lipogenic genes after parturition and through 56 DIM. In contrast, the expression of most of the lipolytic enzymes, receptors and proteins was similar in all cows pre- and postpartum. These results confirm that gene transcription is a major control mechanism f...

BackgroundTranscriptome analysis of porcine whole blood has several applications, which include deciphering genetic mechanisms for host responses to viral infection and vaccination. The abundance of alpha- and beta-globin transcripts in blood, however, impedes the ability to cost-effectively detect transcripts of low abundance. Although protocols exist for reduction of globin transcripts from human and mouse/rat blood, preliminary work demonstrated these are not useful for porcine blood Globin Reduction (GR). Our objectives were to develop a porcine specific GR protocol and to evaluate the GR effects on gene discovery and sequence read coverage in RNA-sequencing (RNA-seq) experiments.ResultsA GR protocol for porcine blood samples was developed using RNase H with antisense oligonucleotides specifically targeting porcine hemoglobin alpha (HBA) and beta (HBB) mRNAs. Whole blood samples (n = 12) collected in Tempus tubes were used for evaluating the efficacy and effects of GR on RNA-seq. The HBA and HBB mRNA transcripts comprised an average of 46.1% of the mapped reads in pre-GR samples, but those reads reduced to an average of 8.9% in post-GR samples. Differential gene expression analysis showed that the expression level of 11,046 genes were increased, whereas 34 genes, excluding HBA and HBB, showed decreased expression after GR (FDR <0.05). An additional 815 genes were detected only in post-GR samples.ConclusionsOur porcine specific GR primers and protocol minimize the number of reads of globin transcripts in whole blood samples and provides increased coverage as well as accuracy and reproducibility of transcriptome analysis. Increased detection of low abundance mRNAs will ensure that studies relying on transcriptome analyses do not miss information that may be vital to the success of the study.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-954) contains supplementary material, which is available to authorized users.