ABSTRACT
Three single nucleotide polymorphisms (SNPs) in the porcine MYOD1 gene were used for association analysis and haplotype construction to evaluate the effects of their substitution. Four hundred and three pigs of Yorkshire and Berkshire breeds were used. The mRNA expression levels of MYOD1 were examined. The g.489C>T and g.1264C>A SNPs were significantly associated with several muscle fiber characteristics, the loin eye area, and lightness. Particularly, animals having hetero-genotypes of both sites showed good performance both in lean meat production and meat quality traits. The results of haplotype substitution were similar to the associations of individual SNPs. Moreover, the 2 SNPs had significant effects on mRNA expression. Therefore, the g.489C>T and g.1264C>A SNPs in MYOD1 may be meaningful DNA markers that can be used for improving important porcine economic traits.
Subject(s)
Genotype , Meat/analysis , Muscle Fibers, Skeletal/metabolism , MyoD Protein/genetics , Phenotype , Polymorphism, Single Nucleotide , Sus scrofa/genetics , Animals , Body Fluid Compartments/metabolism , Breeding , Color , Dietary Fats/metabolism , Meat/standards , MyoD Protein/metabolism , RNA, Messenger/metabolismABSTRACT
The intracellular proteinase of Lactobacillus casei ssp. casei LLG was isolated in the cytoplasmic fraction with 0.05 M Tris-HCl buffer (pH 7.5). The enzyme was purified by the fast protein liquid chromatography system equipped with ion-exchange and gel filtration chromatographies. This proteinase comprised a single monomeric form and had a molecular weight of about 55 kDa and an isoelectric point near pH 4.9. The optimum pH and temperature for the enzyme activity were determined to be pH 6.5 and 37 degrees C, respectively. The enzyme was inactivated by metal-chelating compounds (EDTA, 1,10-phenanthroline) and less affected by serine proteinase inhibitors (diisopropylfluorophosphate, phenylmethylsulfonyl fluoride). Proteinase activity was increased by Ca++, Mn++, and Co++, and inhibited by Cu++, Mg++, and Zn++. The activity of this enzyme to hydrolyze casein appeared to be more active on beta-casein than alphas1-casein and kappa-casein as monitored by polyacrylamide gel electrophoresis.
Subject(s)
Cations, Divalent/pharmacology , Lacticaseibacillus casei/enzymology , Peptide Hydrolases/isolation & purification , Peptide Hydrolases/metabolism , Protease Inhibitors/pharmacology , Cheese/microbiology , Chromatography, Gel , Chromatography, Ion Exchange , Chromatography, Liquid , Electrophoresis, Agar Gel , Hydrogen-Ion Concentration , Hydrolysis , Molecular Weight , Peptide Hydrolases/chemistry , TemperatureABSTRACT
A colorimetric assay for cellular transglutaminase using 5-(biotinamido)pentylamine and polyvinylidine difluoride membranes for crude cellular preparations and purified enzyme has been developed. The biotinpentylamine substrate was incorporated into N,N-dimethylcasein by transglutaminase, the biotinylated products were adsorbed onto the membrane disks and conjugated with streptavidin-beta-galactosidase, and the absorbance resulting from the formation of p-nitrophenol from hydrolysis of p-nitrophenyl-beta-D-galactopyranoside was measured at 405 nm. The validity of the assay was established by showing a good correlation, gamma = 0.922, between the colorimetric procedure and the commonly used radiometric filter paper method for the enzyme. The procedure offers a rapid, sensitive, and nonisotopic method for the estimation of cellular transglutaminase activity in as low as 20 ng of purified guinea pig liver transglutaminase and 10 micrograms of crude fibroblast cytosol protein.
