Wickramasinghe et al. BMC Genomics 2012, 13:45 http://www.biomedcentral.com/1471-2164/13/45 Page 8 of 14 Figure 3 Expression changes in genes involved in milk fat metabolism pathway along the course of lactation. Stages of lactation are shown in x-axis and the RPKM gene expression values are shown in y-axis. Gene names are given in the legend. Fat metabolism in milk was divided in to six sections as: A) Fatty acid uptake, B) De novo fatty acid synthesis, C) Fatty acid desaturation, D) Fatty acid esterification, E) Regulation of fatty acid synthesis, F) Milk fat secretion. Most of the genes in fat metabolism pathway showed high expression in transition and peak lactation milk. acid synthesis, showed a significant decrease in expres- sion along the course of lactation. A similar pattern of expression was observed with the fatty acid synthase (FASN) gene (Figure 3B). Compared to ACACA, the FASN gene showed higher expression throughout lactation. Stearoyl-CoA desaturase (SCD1) is the main enzyme involved in mono-unsaturated fatty acid synthesis in ruminants [42]. SCD1 showed higher expression in tran- sition lactation and decreased expression in peak and late lactation MSC. Enzymes for very long fatty acid desaturation is encoded by the fatty acid desaturase 1 and 2 (FADS1, FADS2) genes. Similar to previous obser- vations in the bovine [25] and rat mammary gland [43], FADS1 showed a higher expression in MSC when com- pared to FADS2. Highest expression of FADS1 was observed in transition lactation, whereas FADS2 showed the highest expression in peak lactation (Figure 3C). Glycerol-3-phosphate O-acyltransferase (GPAM), 1- acylglycerol-3-phosphate O-acyltransferase 6 (AGPAT6) and lipin 1 (LIPIN1) genes encode for the esterification enzymes in mono- and diacylglycerol synthesis. These three genes showed a similar pattern in expression with a high expression in transition lactation MSC and then a progressive decrease in expression along the lactation (Figure 3D). The AGPAT6 gene had the highest expres- sion among these three genes throughout lactation. Dia- cylglycerol transferase 1 and 2 (DGAT1, DGAT2) genes encode for the major enzymes in TAG synthesis. Even though the expression of DGAT2 was low in transition lactation MSC, by the end of lactation the expression of DGAT2 was similar to that of DGAT1 (Figure 3D). Sterol regulatory element-binding proteins 1 and 2 (SREBF1, SREBF2) play an important role in the tran- scription regulation of milk FA synthesis in the mouse [44] and cow [45]. SREBF1 showed a relatively constant expression along the course of lactation whereas SREBF2 had higher expression in peak and late lactation MSC (Figure 3E). Insulin induced 1 and 2 genes (INSIG1, INSIG2) and sterol-sensing proteins (SCAP) are involved in regulating SREBF activity. Both INSIG1 and SCAP had higher expression in peak lactation whereas INSIG2 had high expression in late lactation. Butyrophilin (BTN1A1), Xanthine dehydrogenase (XDH) and adipophilin (ADFP) are main proteins involved in milk fat globule formation [25]. Genes encoding these proteins showed higher expression in transition lactation MSC and a significant decrease in the expression levels for BTN1A1 and XDH1 was observed during the course of lactation (Figure 3F). Expression levels for ADFP remained relatively constant along the course of lactation. Most of the genes involved in lipid metabolism showed high expression in transition lactation MSC. VLDLR and INSIG2 had the highest expression in late lactation milk, whereas FADS2, DGAT1, SREBF2,