| Fat is one of the most important economical traits of pig, and not only affects meat quality, but also is closely related to diseases. We used the Agilent porcine genome commercial microarray to examine the differential gene expression profiling of porcine fat in different body sites (inner layers of backfat, upper layers of backfat and abdominal fat) of Landrace and Rongchang pigs at seven months. The results of array analysis are as follows:(1) Differential expression among each breed, different sex and distinct tissues was identified by T test and variance analysis (ANOVA), and differential expression genes were achieved. The expression changes of these genes may explain molecular mechanism of the distinct differences in fat-deposition ability between Landrace and Rongchang pigs.(2) SOM and HCL analysis revealed that differential expression genes were classified into 9 kinds, involving fat transport, cholesterol metabolism, fatty acid synthesis and other biological process. Gene expression patterns tended to be identical within breeds, and breed effect was more dominant than sex and tissue effect in determining gene expression patterns.(3) Principal constituent analysis (PCA) revealed that differential gene expression levels were higher in Landrace pigs than that of Rongchang pigs. This indicates that it may account for phenotype difference in fat deposition ability between two pigs.(4) GCT based on GO analysis revealed that there are totally 10 GO terms involving in fat metabolism, energy metabolism, cell proliferation and differentiation, and growth was significant (P< 0.05 and 0.01) between genders within each breed and tissue; there were 23 GO terms between two breeds within sex and tissue; 34 GO terms among tissues within each breed and sex, and 27 GO terms between two breeds across gender effect and 3 GO terms on the contrary. These findings suggest that the distinct differences in fat-deposition ability between two breeds may closely correlate with the expression changes of these genes.(5) Gene set enrichment analysis (GSEA) revealed that differential expression genes had significant enrichment in many different pathways including Insulin signaling pathway, Glycolysis/ Gluconeogenesis and Citrate cycle (TCA cycle) which were related to fat metabolism or energy metabolism pathway. The expression patterns (up or down regulation) of these gene sets also were revealed. The potential important genes which appeared many times in different pathways and may play an important role in fat deposition were discovered by leading edge analysis. These expression differences of the potential important genes and the gene sets (pathway) revealed molecular mechanism in fat deposition between the two breeds.(6) Chromosome co-localization analysis revealed that genes related to fat deposition mainly enriched in chromosome 1,2,3,4,6,7,9,13 and 14. These findings suggest that genes regulating fat metabolism existed in these chromosomes mainly. These chromosomes will be more valuable for further study that search the molecular markers of fat deposition. (7) Reliability analysis indicated that the worst coefficient of variation (C.V) in this array was 8.66%, and the best is 3.13%. All C.V of the biological repeats among microarray was less than 10%, The average coefficient of variation (C.V) within Landrace and Rongchang pigs were 5.75%和4.83% respectively. Quantitative PCR was used to verify the microarray data for 10 modulated genes within each tissue, and a good correlation (The highest C.V were 0.950 and 0.974 respectively within Landrace and Rongchang pigs, and the C.V were 0.773 and 0.760 respectively) between the two measures of expression was observed for both two pig breeds. All show the data used in our study is accurate and reliable, and experiment data can reflect the real gene expression patterns.In a word, these results highlight some possible candidate genes and interesting pathways for fat deposition, and provide some data for us to study the molecular mechanism of fat deposition. |
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