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September 3rd, 2010
INTRODUCTION Fat deposition of pigs is of economic importance because of market incentives for lean pork production and decreased feeding costs. It is crucial to investigate and characterize new candidate genes and QTL relevant to pig fat deposit traits. To date, several quantitative trait loci (QTL) significantly affecting 10th-rib, average backfat thickness and other production traits have been mapped on SSC7 (Wang et al., 1998; Nagamine et al., 2003). Peroxisomal [[DELTA].sup.3],[[DELTA].sup.2]-enoyl-CoA isomerase (PECI) was located near the boundary of the quantitative trait loci (QTL) region. [[DELTA].sup.3],[[DELTA].sup.2]-enoyl-CoA isomerase (Ecilp) is unique because its activity is necessary for [beta]-oxidation of all unsaturated fatty acids (Geisbrecht et al., 1999). The series of enzyme-catalyzed reactions required for degradation of fatty acids are evolutionarily conserved and accomplished primarily through the p-oxidation pathway. In peroxisomes, ECI was predicted to be a dominant enzyme for 3-cis 3[right arrow]2-trans and 3-trans 3[right arrow]2-trans isomerizations of long-chain intermediates (Zhang et al., 2002). Fatty acid [beta]-oxidation in mammals is considerably more complicated, primarily due to the existence of overlapping but distinct fatty acid poxidation pathways. Mammalian peroxisomes contain at least three fatty acyl-CoA oxidases, both L-specific and D-specific 2-enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase multifunctional proteins, and at least two thiolases, all of which are encoded by different genes (Palosaari et al., 1990a, 1991; Geisbrecht et al., 1998; Gurvitz et al., 1998; Geisbrecht et al., 1999; Partanen et al., 2004). When the ECI was completely excised in the mouse, it extensively perturbed the metabolism of unsaturated fatty acids, especially for short interval starvation and the fatty acid pattern of complex phospholipids was strongly altered (Palosaari et al., 1990b; Janssen et al., 2002). The PECI gene can be encoded by ECI1 and it is required for growth of saccharomyces cerevisiae on unsaturated fatty acids (Gurvitz et al., 1998). It can be concluded that the PECI gene may play an important role during the metabolic processing of unsaturated fatty acids. Deposition of fat by animals in their bodies is associated with the metabolism of fatty acids, and more research would contribute to understanding of porcine fat deposition. Genomic DNA was isolated from blood of mature Tongcheng pigs (Hubei province, China) by phenol/chloroform extraction. RNA was extracted from muscle tissue of adult Tongcheng pigs and adult Swedish Landrace with TRIzol reagent kit (Life Technologies, Grand Island, NE, USA). RACE (the rapid amplification of cDNA ends) was performed according to the instructions of the SMARTTM RACE cDNA Amplification Kit (Clontech Inc, Palo Alto, CA, USA). The PCR products of RACE were purified with the Wizard PCR Preps DNA Purification System (Promega, Madison, WI, USA). ORF were found by the program SeqMan (DNA star, Madison, WI, USA) and the amino acid sequences were deduced with Primer5.0 (Primer Premier5.0, Premier, Canada). Using the pGEM T-easy vector, DNase I (RNase-free) and M-MLV reverse transcriptase from TaKaRa Dalian (Dalian, China), primers were synthesized (Table 1) and PCR products were sequenced by AuGCT Biotechnology (Bejing, China).
INTRODUCTION Fat deposition of pigs is of economic importance because of market incentives for lean pork production and decreased feeding costs. It is crucial to investigate and characterize new candidate genes and QTL relevant to pig fat deposit traits. To date, several quantitative trait loci (QTL) significantly affecting 10th-rib, average backfat thickness and other production traits have been mapped on SSC7 (Wang et al., 1998; Nagamine et al., 2003). Peroxisomal [[DELTA].sup.3],[[DELTA].sup.2]-enoyl-CoA isomerase (PECI) was located near the boundary of the quantitative trait loci (QTL) region. [[DELTA].sup.3],[[DELTA].sup.2]-enoyl-CoA isomerase (Ecilp) is unique because its activity is necessary for [beta]-oxidation of all unsaturated fatty acids (Geisbrecht et al., 1999). The series of enzyme-catalyzed reactions required for degradation of fatty acids are evolutionarily conserved and accomplished primarily through the p-oxidation pathway. In peroxisomes, ECI was predicted to be a dominant enzyme for 3-cis 3[right arrow]2-trans and 3-trans 3[right arrow]2-trans isomerizations of long-chain intermediates (Zhang et al., 2002). Fatty acid [beta]-oxidation in mammals is considerably more complicated, primarily due to the existence of overlapping but distinct fatty acid poxidation pathways. Mammalian peroxisomes contain at least three fatty acyl-CoA oxidases, both L-specific and D-specific 2-enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase multifunctional proteins, and at least two thiolases, all of which are encoded by different genes (Palosaari et al., 1990a, 1991; Geisbrecht et al., 1998; Gurvitz et al., 1998; Geisbrecht et al., 1999; Partanen et al., 2004). When the ECI was completely excised in the mouse, it extensively perturbed the metabolism of unsaturated fatty acids, especially for short interval starvation and the fatty acid pattern of complex phospholipids was strongly altered (Palosaari et al., 1990b; Janssen et al., 2002). The PECI gene can be encoded by ECI1 and it is required for growth of saccharomyces cerevisiae on unsaturated fatty acids (Gurvitz et al., 1998). It can be concluded that the PECI gene may play an important role during the metabolic processing of unsaturated fatty acids. Deposition of fat by animals in their bodies is associated with the metabolism of fatty acids, and more research would contribute to understanding of porcine fat deposition. Genomic DNA was isolated from blood of mature Tongcheng pigs (Hubei province, China) by phenol/chloroform extraction. RNA was extracted from muscle tissue of adult Tongcheng pigs and adult Swedish Landrace with TRIzol reagent kit (Life Technologies, Grand Island, NE, USA). RACE (the rapid amplification of cDNA ends) was performed according to the instructions of the SMARTTM RACE cDNA Amplification Kit (Clontech Inc, Palo Alto, CA, USA). The PCR products of RACE were purified with the Wizard PCR Preps DNA Purification System (Promega, Madison, WI, USA). ORF were found by the program SeqMan (DNA star, Madison, WI, USA) and the amino acid sequences were deduced with Primer5.0 (Primer Premier5.0, Premier, Canada). Using the pGEM T-easy vector, DNase I (RNase-free) and M-MLV reverse transcriptase from TaKaRa Dalian (Dalian, China), primers were synthesized (Table 1) and PCR products were sequenced by AuGCT Biotechnology (Bejing, China).