The aim of this study was to develop a set of reliable reference genes for quantification of mRNA expression in the pig. The mRNA expression stability in pig tissues was studied for 4 genes: EEF1A1, GAPDH, HPRT1 and TOP2B. The level of expression was characterized by Ct values for each gene and each tissue. By using the geNorm algorithm, the stability of the reference genes was determined in the diaphragm, heart, kidney, liver, lungs, longissimus muscle, and spleen. On the basis of this information, suitable reference genes can be selected for mRNA expression studies in relevant pig tissues.

Skeletal muscle growth is determined by the number of prenatally formed fibres and by the degree of their postnatal hypertrophy; i.e. prenatal development may influence the postnatal growth. Suppression subtractive hybridization (SSH) was used to identify genes more expressed in fetal hind limb muscles of Piétrain pigs (44 days of gestation) compared to the adult biceps femoris. Six potential functional candidate genes (CNN3, DCN, FBN2, POSTN, SPARC and YWHAQ) were selected to verify the SSH results using real-time RT-PCR. Expression levels of the studied genes were significantly higher (P < 0.05) in the fetal muscle compared to the adult muscle. FBN2 and POSTN exhibited the highest mRNA levels (mean relative ratios were 182.7 and 121.6, respectively). The studied genes may play an important role in muscle biology and may be candidates for muscling traits.

Source and description: From the subtracted foetal hind limb muscle cDNA library (44 days of gestation, using adult biceps femoris cDNA as the driver) eight genes (FBN2, YWHAQ, CNN3, DCN, POSTN, SPARC, RBM39 and GNAS) were selected for analysis of polymorphisms and mapping.

Background: Peroxisome proliferator-activated receptor γ coactivator 1α (PPARGC1A) is a coactivator with a vital and central role in fat and energy metabolism. It is considered to be a candidate gene for meat quality in pigs and is involved in the development of obesity and diabetes in humans. How its many functions are regulated, is however still largely unclear. Therefore a transcription profile of PPARGC1A in 32 tissues and 4 embryonic developmental stages in the pig was constructed by screening its cDNA for possible splice variants with exon-spanning primers.Findings: This led to the discovery of 2 new splice variants in the pig, which were subsequently also detected in human tissues. In these variants, exon 8 was either completely or partly (the last 66 bp were conserved) spliced out, potentially coding for a much shorter protein of respectively 337 and 359 amino acids (aa), of which the first 291 aa would be the same compared to the complete protein (796 aa). Conclusion:Considering the functional domains of the PPARGC1A protein, it is very likely these splice variants considerably affect the function of the protein and alternative splicing could be one of the mechanisms by which the diverse functions of PPARGC1A are regulated.

The aim of this study was to verify an effect of RNA storage in different laboratory conditions. Especially, this work was focused on the importance of using diethylpyrocarbonate (DEPC) treated water for storage of isolated RNA. The effect of storage of RNA samples in different temperatures was monitored according to various times as well. Isolated RNA was incubated at 20 °C, 4 °C, −20 °C and −80 °C, whereas the temperature −80 °C was used as a control. After incubation only mRNA was converted to cDNA by reverse transcription. The polymerase chain reaction in real time (real-time PCR) was used for a measurement of RNA degradation. No statistically significant interactions were found between RNA treatment conditions if analysis of variance (ANOVA) model was applied. The result showed that storage of isolated RNA in water treated with DEPC is not necessary. This approach prevents possible inhibition downstream reaction caused by DEPC. The results of this study can be used in all molecular applications based on RNA.