Genetic polymorphism of candidate genes in pig meat production

H-FABP, LEPR and MC5R genes were suggested as candidate genes for fat content in pig meat. The aim of this study was to detect genetic variation in the porcine H-FABP, LEPR and MC5R genes by PCR-RFLP method in a group of pigs. Genotyping of pigs was done by PCR-RFLP methods. We identified three genotypes in the set of pigs, HH (0.504), Hh (0.412) and hh (0.084) for H-FABP (HinfI). Allele H showed higher frequency than allele h (0.710 vs. 0.290). Three genotypes were identified for the H-FABP (HaeIII) gene (DD - 0.194, Dd - 0.494, dd - 0.312). The allele D (0.441) showed slightly lower frequency than allele d (0.559). All three genotypes were identified for LEPR (HpaII) in the group of pigs (AA – 0.137, AB - 0.314, BB – 0.549). Higher frequency of LEPR gene was confirmed for allele B (0.706), as compared with allele A (0.294). We identified two genotypes for MC5R (BsaHI) in the group of pigs (AA - 0.348 and AG - 0.652), genotype GG was not found. As conforms with genotype structure, we recognize a higher frequency of allele A (0.674) as compared with allele G (0.326).


INTRODUCTION
In the process of pig interbreeding, as well as that of other farming animals, the aim of breeders is to achieve the best possible productive results and traits derived from several molecular-genetic methods. Demand for lean pork meat results in breeding programs aimed at fat reduction. Therefore, we have decided to focus on the polymorphism candidate genes which have an effect on pork production, among which we include: H-FABP, LEPR and MC5R.
The H-FABP gene codes for a protein are related to the intracellular transport of fatty acid and play an important role in the regulation of lipid metabolism. Given this physiological role, H-FABP has been considered an interesting candidate gene for intramuscular fat and backfat thickness in pigs. Gerbens et al. (1997). The H-FABP is a protein present in several tissues with a high damage for fatty acids, such as cardiac and skeletal muscle and lactating mammary glands.
Leptin, a product of the LEP gene, is secreted in adipose tissue and signals to the hypothalamus that the body has been saturated. Expression and secretion of leptin is obviously related to food intake regulation, energy metabolism and body weight (Campfield et al., 1995, Remesar et al., 1997. Its receptor (leptin receptor, LEPR) has been localized to several hypothalamic nuclei, including the ventromedial hypothalamus, the medial preoptic area, the paraventricular nucleus, and the arcuate nucleus (Keen-Rhinehart et al. 2004). Vincent (1997) identified a HinfI polymorphism in LEPR gene and mapped its location to pig chromosome 6. Stratil (1998) identified two other types of polymorphism with the restriction enzymes HpaII and RsaI. Emnett et al. (2001) confirmed the LEPR (MboI) genetic effect on average daily gain.
Among candidate genes with an effect on the content of fat in pork belongs gene of melanocortic receptor MC5R. Gene MC5R belongs to the family of melanocorticotrophic genes and coding the systems of receptor for adenocorticotrophic hormone. Kim et al. 2000 localized MC5R gene to pigs 6 chromosome. Kim et al., 2000 detected two types of MC5R gene polymorphism by using restriction enzyme BsaHI with splitting polymorphic fragments A (238 bp) and G (179 bp and 59 bp). Second type of polymorphism detected by specific PCR shows alleles C (128 bp) and T (118 bp). Protein MC5R is a mediator for thermoregulation, secretion of glands and sexual behavior (Kim et al., 2000). The analysis of a whole population shows the significant influence of the MC5R (BsaHI) gene on backfat thickness where the allele A was in association with higher backfat thickness (Emnett et al., 2001).
Application of these candidate genes direct in selection programs give possible way to obtain decrease backfat thickness, without reduction of intramuscular fat.

RESULTS and DISCUSION
Out of the PCR-RFLP analysis results were calculated frequencies of genotypes and alleles. Genetic structure of observed genes is presented in Table 3. Effectiveness of the alleles was evaluated and presented in Table 4.

LEPR (HpaII)
All three genotypes were identified for LEPR in the group of pigs (AA -0.137, AB -0.314, BB -0.549). Higher frequency of LEPR gene was confirmed for allele B (0.706) against allele A (0.294). Stratil et al. (1998) reported similar results, where the B allele was the predominant in the Landrace, Meishan, Large White and Czech meat pig. Amills et al. (2008) confirmed a high occurrence of pigs with genotype BB (0.659) in the breed Landrace.

MC5R (BsaHI)
We identified two genotypes in the group of pigs (AA -0.348 and AG -0.652); genotype GG was not found. In conformity with genotype structure, we recognize a higher frequency of allele A(0.674) compared with allele G(0.326). The highest occurrence individuals in the set have genotype AG (0,652) while Kim et al. (2000) and Emnett et al. (2001) present abundance of homozygotes AA (Table 3-4.).

CONCLUSIONS
The genotypes of the leptin receptor (LEPR -HpaII), MC5R (BsahI) and cardio-fatty acid binding protein (H-FABP -HinfI, HaeIII) genes were determined for the hybrid pigs. The PCR-RFLP methods were used for genotyping. These genes were suggested as candidate genes for fat content in pig meat.

ACKNOWLEDGEMENT
This article was part of the project VEGA 1/0061/10