An Investigation into the Genetic
Controls of Pork Quality
Rebecca Emnett1,
Steven Moeller1, Keith Irvin1,
1The Ohio State University, Department of Animal Sciences, Columbus, Ohio Introduction Consumers and many sectors of the pork industry are demanding improvements in meat quality. This provides a new challenge for the breeding industry, which is seeking advanced genetic tools that can be practically incorporated in selection schemes for trait improvement. The heightened interest in meat (muscle) quality has led to investigations into the physiological and genetic controls of these economically important traits. Often studies in livestock are modeled after known rodent or human genetic markers associated with diseases, which may also be related to body composition or changes in energy metabolism. Genetic markers characterized in other species serve as guides or “candidate” genes for investigation of the phenotypic differences found in swine. Although several genetic markers
and QTLs affecting meat quality and performance traits have been detected in
the pig (for review see Rothschild and Plastow, 1999), the quest continues to
identify markers that explain significant variation in these traits of economic
importance. It is anticipated that the detection of new molecular genetic
markers, together with advances in the area of quantitative genetics, will lead
to the development of marker assisted selection (MAS) programs for meat quality
improvement and practical utilization by swine producers. As more information becomes
available in swine molecular genetics, there exists a need to analyze
associations between these markers and phenotypes in individual genetic lines
or populations. The objective of this study was to determine the association
between variation found in performance, carcass, and meat quality traits and
several candidate genes of interest in different pig populations. Materials & Methods
Candidate Gene Association
Studies The population utilized for the
project consisted of both purebred and crossbred animals from the 1998 National
Barrow Show Progeny Test and the 1999 Hampshire Sire Progeny Test. All animals
were managed by identical protocols, and performance, carcass, meat quality,
and sensory panel
measurements were taken on each animal.
Individuals of four major U.S. breeds were chosen for DNA extraction
from frozen loin chops. Berkshire (n=180), Duroc (n=77), Hampshire (n=160), and
Landrace (n=55) sire breeds were represented. Genotypes were obtained for six
candidate genes using Polymerase Chain Reaction – Restriction Fragment Length
Polymorphism (PCR-RFLP) procedures unique for each gene analyzed. Statistical
analyses were performed within each breed separately, and across breeds for the
total population. Only pigs classified as free of the Porcine Stress Syndrome
mutation were utilized in these analyses. A SAS (1999) Mixed Model analysis was
completed for the total population utilizing the fixed effects of breed, genotype, sex, day off-test group
(depending on the trait), and Rendement Napole (RN) gene status, with a
sire(breed) random effect. The within breed analysis included a random effect
of sire, and fixed effects of genotype, sex, day off-test group (depending on
the trait), and RN classification. A second PPARg analysis excluding the underrepresented 22-genotype animals
(n=12) was performed to more clearly define the differences between the 11 and
12-animals. Allelic frequencies were calculated within breed and for the total
population based upon observed genotype classifications. Mapping Procedures Two additional genes were chosen
as possible candidate genes for meat quality based on their observed
physiological functions in other mammalian species. Primers for use in PCR
amplification were designed based on porcine sequence available for each gene
in GenBank. PCR conditions were optimized for each gene separately. Sequencing
and comparisons of breed DNA pools were completed in order to search for
sequence variation among the breeds. Physical mapping was achieved by use of a
pig-rodent somatic cell hybrid panel (Yerle et al., 1996). Results Leptin Receptor (LEPR)
Daily gain and backfat thickness are important traits
for livestock producers to consider in order to produce efficient, fast growing
and lean animals. The leptin receptor gene (LEPR),
is a high affinity receptor (for review see Tartaglia, 1997) that mediates the
regulation of the well known “obesity” gene, leptin (Zhang et al., 1997).
Mutations in LEPR have been reported
to be associated with obesity in humans and rodents (Reichart et al., 2000;
Clement et al., 1998; Chen et al., 1996). Given the physiological role of LEPR, it is interesting as a candidate
gene for backfat deposition and daily gain in the pig. Vincent et al. (1997) identified a HinfI polymorphism in porcine LEPR
and mapped its location to pig chromosome 6 (SSC6). A few studies have reported
on associations between leptin levels and
production traits in pigs (Ramsay et al. 1998; Robert et al. 1998), but the
effects of LEPR on pork quality
traits of economic importance to the industry have not been investigated. A leptin receptor (LEPR) MboI RFLP,
developed by Vincent at Iowa State University (M.F. Rothschild, personal
communication), was found to be polymorphic in all breeds analyzed; Berkshire
(n=177), Duroc (n=76), Landrace (n=49), and Hampshire (n=149). Allele 2 was the
most frequent (.91) in the total population and was found to be associated with
leaner animals (Table 1). Total population analysis revealed effects (P<.05) of LEPR on last lumbar backfat with the 11-animals having the fattest
phenotype (Table 1). Although not significant (P>.05), last rib and 10th rib backfat (not shown) had
similar numerical trends. Average daily gain was also different (P<.05) between the genotypes
(Table 1). The results of the individual breed analyses (not shown) revealed
differences (P<.05) between
LEPR genotypes and off flavor score for
Berkshire; average daily gain and Japanese color score for Duroc; glycolytic
potential, loin glycogen and lactate concentration, intramuscular fat, and
quality index (which includes Minolta, IMF, & pHu) for
Hampshire; 10th rib backfat, average daily gain, loin pHu,
Minolta and Hunter color values, and quality index for Landrace. As previously reported in other mammalian species, LEPR appears to have the greatest
effects on backfat and average daily gain in the pig, but may also be
associated with correlated pork quality differences within genetic lines.
Melanocortin-4 Receptor (MC4R) Melanocortin-4 receptor (MC4R) has been found to play a significant role in regulating leptin’s effects on food intake and body weight (Seeley et al., 1997; Fan et al., 1997). Kim et al. (2000) demonstrated that a missense mutation in MC4R was associated with backfat thickness, growth, and feed intake in different genetic lines of pigs. No studies to date have reported associations between meat quality characteristics and MC4R genotypes. Results with the TaqI
MC4R RFLP, developed by Kim et al. (2000),
show an allelic frequency of .60 for allele 2, which was associated with much
fatter animals in the total population. Genotypic frequencies varied within the
breeds, however the heterozygote 12-animals were the most frequent (.45) in the
total population. The effect of MC4R on last lumbar backfat was highly significant (P<.001) (Table 1). Approximately, 0.18 cm in last lumbar backfat is added with the inclusion of each 2 allele. MC4R genotype groups were also different (P<.05) for last rib backfat. and 10th rib backfat approached significance (P=.085) with the fatter 22-animals being in line with the last lumbar result. These backfat results are also in agreement with Kim et al. (2000) who demonstrated that 11-homozygote pigs had approximately 9 % less backfat than 22-pigs. Instron force was less (P<.05) for the 22-genotype (Table 1), indicating increased tenderness in the fatter animals. Interesting trends in loin muscle area, average daily gain, and color score were also noted. The results of the individual breed analysis (not shown) revealed differences (P<.05) between MC4R genotypes and soundness score, last lumbar backfat, last rib backfat, average backfat, average daily gain, and loin lactate concentration for Berkshire; and juiciness score, intramuscular fat, Minolta, Hunter color, and quality index for Landrace. Our results suggest that MC4R has its greatest effects on backfat and Instron tenderness for the total population. However, differences between MC4R genotypes were also reported for average daily gain and meat quality traits within the breed populations. Further analysis in larger populations will be needed with this marker to fully characterize the effects of MC4R on meat quality. Mealnocortin-5 Receptor (MC5R) Melanocortin-5 receptor (MC5R) has been found to be associated
with thermoregulation through gland secretion (van der Kraan et al., 1998; Chen
et al., 1997) and serves a possible role in lipolysis of adipocytes (Boston,
1999). Kim et al. (1999) mapped porcine MC5R
to SSC 6 and detected two single nucleotide polymorphisms within the porcine
sequence. Previous analyses have not investigated the association between the
porcine MC5R gene and fat deposition,
growth or carcass quality traits in pigs. Results indicate that the MC5R BsaHI RFLP, developed by Kim et al. (1999) was polymorphic in
Berkshire, Duroc, Landrace and Hampshire populations. The frequency of alelle 1
was .82. Genotypic frequencies varied within the breeds, however, the
homozygote 11-animals were the most frequent (.75) in the total population. Total population analysis revealed effects (P<.05) of MC5R on 10th rib backfat (Table 1), with the 11 and
12-animals being fatter. Similar numerical differences were also noted for the
last rib location (P=.14). Loin
muscle area also approached significance for the total population analysis (P=.10). The results of the individual
breed analysis (not shown) revealed differences (P<.05) between MC5R
genotypes with color and Japanese subjective scores, Minolta and Hunter color
values, as well as, Instron tenderness for Berkshire; quality index for
Hampshires; all backfat measures and intramuscular fat % for Landrace. This MC5R
BsaHI RFLP appears to have its greatest effects on backfat in the total
population. Further analysis is needed to fully characterize the differences in
meat quality characteristics such as color and intramuscular fat % within the
breed populations. Peroxisome Proliferator
Activated Receptor-g (PPARg) Peroxisome Proliferator Activated Receptor-gamma (PPARg) is a member of the nuclear receptor superfamily (for review see Green, 1995) and regulates the expression of several genes encoding proteins involved in adipocyte differentiation (Rosen et al., 2000; Spiegelman et al., 1997) and fat deposition (for review see Schoonjans et al., 1996). Genetic mutations in PPARg have been found to be associated with extreme obesity in humans (Freake, 1999). In pigs, PPARg expression levels in adipose tissue vary among different breeds and ages (Grindflek et al., 1998). An association study (Grindflek, in manuscript), reported a difference in loin fatty acid composition in Norwegian pigs for a BsrI RFLP, but no significant differences were noted for backfat or intramuscular fat measurements. Our results show that a BsrI PPARg RFLP
(Grindflek, personal communication) was polymorphic in Berkshire, Duroc,
Landrace, and Hampshire populations. The frequency of allele 1 was .81. Total
population analysis revealed effects (P<.05)
on off flavor score (Table 1), although interesting trends in average daily
gain, tenderness and juiciness approached significance. The results of the
individual breed analysis (not shown) revealed differences (P<.05) between PPARg genotypes and loin muscle
area and marbling for Duroc, average daily gain for Hampshire; and last rib
backfat, and Instron tenderness for Landrace.
This PPARg RFLP remains an interesting
candidate gene for meat quality traits within specific lines of swine. These results,
while promising, warrant larger scale investigation to determine the potential
use of PPARg in
future selection programs. Heart Fatty Acid Binding
Protein (HFABP) Intramuscular fat percentage (IMF) has been found to be positively associated with sensory attributes of pork (Fernandez et al., 1999; Touraille et al., 1989). Hovenier et al. (1992) reported that backfat reduction is not completely related with reductions in IMF. Therefore, it may be possible to treat the two traits separately in a breeding scheme with the proper selection tools. Heart Fatty Acid Binding Protein-1 (HFABP) is a member of the fatty acid binding protein family (FABP), which is involved in fatty acid transport from the cell membrane to the intracellular sites of fatty acid utilization (Veerkamp and Maatman, 1995). Given this physiological role, HFABP has been considered to be an interesting candidate gene for IMF and backfat in pigs. Gerbens et al. (1997) mapped HFABP to pig chromosome 6. QTL studies have also identified IMF and BF loci in this region of chromosome 6 (Ovilo et al. 2000b; de Koning et al., 1999), further implicating HFABP as a strong candidate gene for IMF and backfat in the pig. Gerbens et al. (1997) reported three polymorphic sites in the porcine HFABP gene (HaeIII, MspI, and HinfI) and conducted an association study to determine the genotype effects on traits in pigs (Gerbens et al., 1999). This study reported IMF and backfat differences between HFABP genotype groups and thus hypothesized that HFABP, or a closely linked marker, controlled IMF differences in pigs. Two of these markers were also informative in a Norwegian pig population (E. Grindflek, personal communication), and Ovilo et al. (2000a) also reported differences in genotype groups for the HFABP HaeIII RFLP. Results from our population show that the HaeIII HFABP RFLP (originally reported by Gerbens et al., 1997) is
polymorphic in Berkshire, Duroc, Landrace and Hampshire populations. The
frequency of allele 1 was .63. Total population analysis reveals effects of HFABP on ultimate loin pH (P<.05) and quality index (P<.01) (Table 2), and
intramuscular fat % and pork flavor also showed interesting trends. The results
of the individual breed analysis (not shown) revealed differences between HFABP genotypes and loin muscle area,
loin glycogen concentration, intramuscular fat, Instron tenderness, and quality
index for Berkshire. The Berkshire analysis results are similar to those of
Gerbens et al. (1999), which found an advantage of the HaeIII HFABP 12-heterozygote
for intramuscular fat % and backfat.
Loin glycogen concentration was also significantly different (P<.05) in Duroc; last rib
backfat, flavor score, and water holding capacity for Hampshire; and last rib
backfat, and intramuscular fat for Landrace. HFABP remains an interesting candidate gene for meat quality traits
of importance within specific breeds of swine.
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