A porcine gene, PBK, differentially expressed in the longissimus muscle from Meishan and Large White pig

An investigation of differences in gene expression in the longissimus muscle of Meishan and Large White pigs was undertaken, using the mRNA display technique. A fragment of one differentially expressed gene was isolated and sequenced, whereupon the complete cDNA sequence was then obtained by using the rapid amplification of cDNA ends (RACE). The nucleotide sequence of the gene is not related to any known porcine gene. Sequence analysis revealed that the open reading frame of this gene encodes a protein with 322 amino acids, thus displaying high sequence identity with the PDZ binding kinase (PBK) of eleven other animal species - dog, horse, cattle, human, chimpanzee, crab-eating macaque, rhesus monkey, rat, mouse, gray short-tailed opossum and platypus, so it can be defined as the porcine PBK gene. This gene was finally assigned GeneID:100141310. Phylogenetic tree analysis revealed that the swine PBK gene has a closer genetic relationship with the PBK gene of platypus. Gene expression analysis of eight tissues of a Meishan x Large White cross showed that the porcine PBK gene is differentially expressed in various tissues. Our experiment established the primary foundation for further research on this gene.     

Molecular characterization and expression profile of a novel porcine gene differentially expressed in the muscle and backfat tissues from Chinese Meishan and Russian Large White pigs

The mRNA differential display technique was performed to investigate the differences of gene expression in the longissimus dorsi muscle and backfat tissues from Chinese Meishan and Russian Large White pigs. One novel gene that was differentially expressed was identified through semiquantitative RT-PCR, and the cDNA complete sequence was then obtained using the rapid amplification of the cDNA ends (RACE) method. The cDNA sequence of this gene is not homologous to any of the known porcine genes. The sequence prediction analysis revealed that the open reading frame of this gene encodes a protein of 402 amino acids that contains the putative conserved transposase DDE domain, and further Blast analysis revealed that this protein has 100% homology with the Tn10 transposase from Oryza sativa, Serratia marcescens, and Salmonella, and, therefore, this gene can be defined as the swine Tn10 transposase gene. This novel porcine gene was finally assigned to Gene ID: 100049649. The RT-PCR analysis of the tissue expression profile was carried out using the tissue cDNAs of one Meishan pig as the templates, and the result indicated that this novel swine gene is moderately expressed in fat and weakly expressed in small intestine, liver, kidney, and spleen but almost not expressed in heart, ovary, muscle, and lung. Our experiment established the primary foundation for further research into the biological significance of swine Tn10 transposase gene.     

Molecular characterization and expression profile of a novel porcine gene differentially expressed in the muscle and backfat tissues from Chinese Meishan and Russian Large White pigs

The mRNA differential display technique was performed to investigate the differences of gene expression in the longissimus dorsi muscle and backfat tissues from Chinese Meishan and Russian Large White pigs. One novel gene that was differentially expressed was identified through semi-quantitative RT-PCR and the cDNA complete sequence was then obtained using the rapid amplification of cDNA ends (RACE) method. The cDNA sequence of this gene is not homologous to any of the known porcine genes. The sequence prediction analysis revealed that the open reading frame of this gene encodes a protein of 402 amino acids that contains the putative conserved transposase DDE domain and further Blast analysis revealed that this protein has 100% homology with the Tn10 transposase from Oryza sativa, Serratia marcescens, and Salmonella, and therefore, this gene can be defined as the swine Tn10 transposase gene. This novel porcine gene was finally assigned to Gene ID: 100049649. The RT-PCR analysis of the tissue expression profile was carried out using the tissue cDNAs of one Meishan pig as the templates, and the result indicated that this novel swine gene is moderately expressed in fat, and weakly expressed in small intestine, liver, kidney, and spleen but almost not expressed in heart, ovary, muscle, and lung. Our experiment established the primary foundation for further research into the biological significance of swine Tn10 transposase gene.     

Differential display reveals a novel pig gene,PRPF3, which is differentially expressed in Large White versus Wujin skeletal muscle tissues

The mRNA differential display technique was performed to investigate gene expression differences in the longissimus dorsi muscle from Wujin and Large White pigs. A fragment of one differentially expressed gene was isolated and sequenced. A complete cDNA sequence of the gene was obtained using the rapid amplification of cDNA ends method. The open reading frame of this gene encodes a protein of 683 amino acids, which is homologous with the PRP3 pre-mRNA processing factor 3 (PRPF3) of five species: bovine (99%), human (99%), sumatran orangutan (99%), mouse (99%) and chicken (94%). This newly identified gene was respectively defined as the swine PRPF3 gene. This gene is structured in 15 exons and 14 introns as revealed by computer-assisted analysis. The phylogenetic tree analysis revealed that the swine PRPF3 gene has a closer genetic relationship with the PRPF3 gene of sumatran orangutan.     

Molecular cloning,polymorphism and association analyses of a novel porcine mRNA differentially expressed in the Longissimus muscle tissues from Meishan and Large White pigs

The mRNA differential display technique was performed to investigate the differences of gene expression in the longissimus muscle tissues from Meishan and Large White pigs. One novel mRNA that was differentially expressed was identified through semi-quantitative RT-PCR and the cDNA complete sequence was then obtained using the rapid amplification of cDNA ends (RACE) method. The nucleotide sequence of the mRNA is not homologous to any of the known porcine genes. Sequence prediction analysis revealed that the this mRNA is not protein-coding mRNA. Polymorphism analyses revealed that there was a C-T mutation on the position of 669 bp and PCR -Dra I-RFLP analyses revealed that Chinese indigenous pig breeds and exotic pig breeds displayed obvious genotype and allele frequency differences at this locus. Association analyses revealed that this polymorphic locus was significantly associated with the drip loss rate, skin percentage, meat color value (m.Longissimus Dorsi, LD), loin eye width, loin eye area, water holding capacity, carcass length, caul fat weight, intramuscular fat (m.Longissimus Dorsi, LD), lean meat weight, lean meat percentage, backfat thickness at buttock (P < 0.05).     

Detection of differentially expressed genes in the longissimus dorsi of Northeastern Indigenous and Large White pigs

Recent attention in pig breeding programs has focused on the improvement of pork quality in response to increasing consumer demands. Compared to the fatty-type Northeastern Indigenous (Chinese) breed of pigs, the lean-type Large White has lower intramuscular fat and inferior eating quality from the perspective of the Chinese consumer. In order to investigate the molecular basis of differences in pork quality in Chinese indigenous and Western breeds, longissimus dorsi samples were collected from three adult Northeastern Indigenous and three adult Large White pigs. The RNAs were extracted and hybridized to the porcine Affymetrix GeneChip. Microarray analysis demonstrated differential expression of 1134 genes of which 401 have a known function. One hundred and thirty-six genes were up-regulated and 998 down-regulated in Northeastern Indigenous breed compared to Large White pigs. We screened 10 genes as candidate genes associated with pork quality. We investigated a single nucleotide polymorphism in the 5' regulatory region of the gene FABP4 in 65 Songliao black swine, using PCR-single-strand conformational polymorphism. We found this polymorphism to be highly significantly associated with marbling and intra-muscular fat content (P ≤ 0.01). Genotype BB had higher marbling than AB and AA, but there was no significant difference between AB and AA. Genotype BB and AB had higher intra-muscular fat content than AA, but there was no significant difference between BB and AB. These results help to elucidate the genetic mechanisms behind differences in pork quality and provide a theoretical basis for selection and genetic improvement of meat quality traits in pigs.     

A novel porcine gene,MAPKAPK3, is differentially expressed in the pituitary gland from mini-type Diannan small-ear pigs and large-type Diannan small-ear pigs

The mRNA differential display technique was performed to investigate the differences of gene expression in the pituitary gland from mini-type Diannan small-ear pigs and large-type Diannan small-ear pigs. One novel gene differentially expressed was identified through semi-quantitative RT-PCR and its cDNA complete sequence was then obtained using the rapid amplification of cDNA ends (RACE) method. Nucleotide sequence of the gene is not homologous to any of the known porcine genes. The sequence analysis revealed that the open reading frame of this gene encodes a protein of 384 amino acids has high homology with the mitogen-activated protein kinase-activated protein kinase 3 (MAPKAPK3) of eight species-cattle (96%), horse (96%), rhesus monkey (93%), rabbit (77%), human (98%), chimpanzee (98%), mouse (94%) and rat (91%)-so that it can be defined as swine MAPKAPK3 gene. This gene was finally assigned GENE ID: 100233192. Computer assisted analysis revealed that the genomic DNA of open reading frame of the swine MAPKAPK3 gene consists of ten exons and nine introns. The phylogenetic tree analysis revealed that the swine MAPKAPK3 gene has a closer genetic relationship with the MAPKAPK3 of cattle. The tissue expression analysis indicated that the MAPKAPK3 mRNA expression levels of large-type Diannan small-ear pigs are ubiquitously increased in various tissues compared to the mini-type Diannan small-ear pigs. Our experiment suggested that the swine MAPKAPK3 gene might play an important role in the growth and development differences between mini-type Diannan small-ear pigs and large-type Diannan small-ear pigs.     

Identification and characterization of a differentially expressed protein (CAPZB) in skeletal muscle between Meishan and Large White pigs

Actin capping protein beta (CAPZB) protein was identified with considerable differences in the longissimus dorsi muscle between Large White and Meishan pigs using proteomics approach. However, in pigs, the information on CAPZB is very limited. In this study, we cloned and characterized the porcine actin capping protein beta (CAPZB) gene. In addition, we present two novel porcine CAPZB splice variants CAPZB1 and CAPZB2. CAPZB1 was expressed in all twenty tissues. However, CAPZB2 was predominantly expressed in the skeletal muscle and heart. In addition, the two isoforms had different expression profiles during the skeletal muscle development and between breeds. Moreover, the SNP T394G was identified in the coding region of the CAPZB gene, which was significantly associated with the carcass traits including the LFW, CFW, SFT and LEA. Data presented in our study suggests that the CAPZB gene may be a candidate gene of meat production trait and provides useful information for further studies on its roles in porcine skeletal muscle.     

Meat quality of the longissimus lumborum muscle of Casertana and Large White pigs: metabolomics and proteomics intertwined

Longissimus lumborum muscles from high fat-deposing Casertana and lean meat Large White pigs were assayed for meat quality parameters, including early and ultimate post mortem pH, water holding capacity and Minolta L*a*b*values. These parameters were correlated to results from differential proteomic and targeted metabolomic analyses. Higher levels of glycolytic enzymes and lactate accumulation were related to slow pH drop in Casertana pigs, albeit not to rapid pH lowering in LW counterparts. On the other hand, the individuation of pyruvate kinaseM1 and tropomyosin levels in LW were related to water holding capacity and Minolta values at 24h after slaughter. Bioinformatic analyses strengthened the correlation between over-expression of structural proteins in LW and more accentuated growth aptitude in this breed. Conversely, enzymes taking part into branching glycolytic reactions, such as glycerol 3-phosphate and creatine kinase M, were related to accentuated lipogenesis and slower albeit prolonged glycolytic rate in Casertana, respectively. Breed-specific differences at the protein level were not only related to growth performances and fat accumulation tendency in vivo, but they also affected post mortem performances through a direct influence on the forcedly anaerobic behavior of pig muscles after slaughter.     

Genetic parameters for tissue and fatty acid composition of backfat, perirenal fat and longissimus muscle in Large White and Landrace pigs

Genetic parameters pertaining to the same chemical characteristics of three porcine tissues, that is backfat (BF), perirenal fat (PF) and longissimus muscle (LM), were estimated in centrally tested Large White and Landrace pigs. Animals were fed ad libitum. They were slaughtered at an average BW of 99.6 kg, and samples of BF (both inner and outer layers) and LM were removed at the 13th to 14th rib level of the carcass on the day after slaughter. The data set included 2483 animals recorded for average daily gain (ADG; 35 to 100 kg), estimated carcass lean percentage (LEAN) and lean tissue growth rate (LTGR). Among these animals, around 950 pigs were recorded for lipid content (L%) and water content (W%) of BF and LM and for fatty acid composition (FAC) of BF, whereas FAC of LM was measured on 297 pigs and L%, W%, and FAC of PF on around 210 pigs. Heritabilities (h2) and genetic correlations (ra) were estimated using REML-animal model methodology. Estimates of h2 for L%, W% and FAC of BF, PF and LM were of moderate-to-high magnitude: for example 0.47 ± 0.09 for L% of LM, 0.59 ± 0.11 for W% of BF, 0.45 ± 0.08 for the ratio of polyunsaturated to saturated fatty acids (P/S) of BF, 0.61 ± 0.15 and 0.29 ± 0.10 for the coefficient of unsaturation of lipids (UNSAT, average number of double bonds of unsaturated fatty acids) of PF and LM, respectively. Genetic correlations of L% with P/S or UNSAT were strongly negative (from -0.4 to -0.9) in BF and LM, but not in PF. The 'between-tissue' genetic correlations for homologous compositional traits were far from being unity (e.g. ra = 0.57 ± 0.05 'between' BF and PF for UNSAT). Genetic relationships between ADG and tissue compositional traits were globally weak. By contrast, genetic correlations were moderate-to-high between carcass leanness and tissue compositional traits, especially those of fat depots: for example -0.66 ± 0.14 between LEAN and L% of BF, 0.50 ± 0.07 between LEAN and UNSAT of PF, -0.44 ± 0.08 between LEAN and L% of LM, and 0.27 ± 0.03 between LEAN and UNSAT of LM. On the basis of the parameter estimates found here, breeding for higher LTGR is expected to increase the ratio of water to lipids and the unsaturation degree of lipids in subcutaneous BF and, to a lesser extent, in PF. Tissue composition and FAC of LM would be less affected.     

Identification of a differentially expressed gene PPP1CB between porcine Longissimus dorsi of Meishan and Large WhitexMeishan hybrids

To study the molecular basis of heterosis, suppression subtractive hybridization was used to investigate the differences in gene expression between porcine Longissimus dorsi of F1 hybrids Large WhitexMeishan and their female parents Meishan. From two specific subtractive cDNA libraries, the clones selected by reverse Northern high-density blot screening were chosen to clone full-length cDNA by rapid amplification of cDNA ends. An expression-upregulated gene for Meishan skeletal muscle, designated protein phosphatase 1, catalytic subunit, beta isoform (PPP1CB), was identified. Porcine PPP1CB contains an open reading frame encoding 327 amino acid residues with 13 and 1763 nucleotides in the 5' and 3' untranslated regions, respectively. A DNA fragment of 721 nucleotides was amplified and a mutation that creates/disrupts a restriction site for endonuclease RsaI was found. The derived amino acid sequence of PPP1CB has high homology with the PPP1CB of three species, Mus musculus (99%), human (99%) and mouse (100%). The tissue expression analysis indicated that the swine PPP1CB gene is generally expressed in most tissues. The possible role of PPP1CB and its relation to porcine heterosis are discussed.     

A novel sheep gene,MMP7, differentially expressed in muscles from black-boned sheep and local common sheep

Differences in gene expression in muscles from Chinese black-boned sheep and local common sheep were investigated using mRNA differential display. One differentially expressed novel gene was identified through semi-quantitative RT-PCR, and the full-length cDNA sequence was then obtained using the rapid amplification of cDNA ends (RACE). The nucleotide sequence of this gene is not homologous to any of the known sheep genes, but it contains an open reading frame encoding a protein of 416 amino acids, which has high homology with matrix metallopeptidase 7 (matrilysin, uterine) (MMP7) of 10 species: bovine (93%), rhesus monkey (75%), human (74%), pig (73%), chimpanzee (73%), dog (73%), horse (72%), mouse (66%), rat (65%), and chicken (53%). Thus the novel gene can be defined as the sheepMMP7 gene. It was finally assigned to GeneID: 100192317. The phylogenetic tree analysis revealed that the sheepMMP7 gene is closely related to the bovineMMP7. Our experiment is the first one to establish the primary foundation for further research on the sheepMMP7 gene.