The partial 16 S r RNA gene sequences（100 to 500 bp） were widely used to reveal rumen bacterial composition influenced by diets, while quantification of the changed uncultured bacteria was inconvenient due to diffic...The partial 16 S r RNA gene sequences（100 to 500 bp） were widely used to reveal rumen bacterial composition influenced by diets, while quantification of the changed uncultured bacteria was inconvenient due to difficult designing of specific primers based on short sequences. This study evaluated the effect of forage resources on rumen bacterial diversity and developed new strategy for primer design based on short sequences to quantify the changed uncultured bacteria. Denaturing gradient gel electrophoresis（DGGE） analysis and subsequent band sequencing were used to reveal the distinct rumen bacteria composition in cows fed with two forage sources（single corn stover vs. mixed forages including alfalfa hay and corn silage）. The bacterial diversity in the rumen of dairy cows fed with corn stover was lower than that with mixed forages（P0.05）. The bacterium named R-UB affiliating to uncultured Succinivibrionaceae was identified, and it was abundant in the rumen of cows fed with mixed forages compared to corn stover. The full length 16 S r RNA gene sequences with identity of 97% to the R-UB 16 S r RNA gene sequence were obtained from Gen Bank and used to design specific primers to quantify uncultured bacterium R-UB. All sequences of amplicon from the new primers were of 100% identity to R-UB sequences indicating the high specificity of new primers. Quantitative PCR confirmed that abundance of R-UB in the rumen of cows fed with corn stover was lower than those fed with mixed forages（P0.01）. New strategy for designing primers based on partial 16 S r RNA genes to quantify targeted uncultured bacteria was successfully developed. The rumen bacteria descending significantly in the cows fed corn stover compared to those fed mixed forages was identified as uncultured R-UB from Succinivibrionaceae.展开更多
Dairy cows undergo tremendous changes in physiological, metabolism and the immune function from pregnancy to lac- tation that are associated with cows being susceptible to metabolic and infectious diseases. The object...Dairy cows undergo tremendous changes in physiological, metabolism and the immune function from pregnancy to lac- tation that are associated with cows being susceptible to metabolic and infectious diseases. The objective of this study is to investigate the changes of plasma proteome on 21 d before expected calving and 1 d after calving from dairy cows using an integrated proteomic approach consisting of minor abundance protein enrichment by ProteoMiner beads, protein labeling by isobaric tags for relative and absolute quantification, and protein identification by liquid chromatography coupled with tandem mass spectrometry. Nineteen proteins were changed around the time of calving. These proteins were asso- ciated with response to stress, including acute-phase response and defense response, based on the proteins annotation. In particular, three up-regulated proteins after calving including factor V, a2-antiplasmin and prothrombin were assigned into the complement and coagulation pathway. These results may provide new information in elucidating host response to lactation and parturition stress, and inflammatory-like conditions at the protein level. Differential proteins may serve as potential markers to regulate the lactation and parturition stress in periparturient dairy cows.展开更多
A PCR method for detection of bovis, sheep, pig, and chicken derived materials in feedstuff was established, and the existing method was improved according to the research on general primer and species-specific primer...A PCR method for detection of bovis, sheep, pig, and chicken derived materials in feedstuff was established, and the existing method was improved according to the research on general primer and species-specific primers. First, general primer designed according to 16S rRNA gene sequence of hovis, sheep, pig, chicken, fish, and horse mtDNA was used for primary detection of animal derived materials in feedstuff. Species-specific primers designed according to conserved sequence of mtDNA of bovis, sheep, pig, and chicken were used for amplification of a 271,274, 149, and 266 bp fragment, respectively. Further confirmation of the detection result was then carried out. PCR method for detection of animal derived materials in unknown feedstuff was developed by using general primer, relevant PCR system, and PCR condition. Also a PCR method for detection of each species （bovines, sheep, pig, and chicken） was designed by using our speciesspecific primers. High sensitivity and specificity of our method were confirmed with a minimum detection level of 0.1%. Method for detection of animal derived materials in this research is not only cheap and easy for operation but also precise and reliable results can be obtained. It could be one of the effective methods for the detection of animal derived materials in feedstuff.展开更多
To determine the effect of dietary supplementation with vegetable oilseeds on the composition of bovine milk fatty acids （FAs）, 40 Holstein dairy cows were used with a complete randomized design. At the beginning of...To determine the effect of dietary supplementation with vegetable oilseeds on the composition of bovine milk fatty acids （FAs）, 40 Holstein dairy cows were used with a complete randomized design. At the beginning of the experiment, the cows were 150±25 day in milk （DIM）. Total duration of the experiment was six weeks. Measurements were made during the last three weeks. Cows in four treatments were fed with a basal diet （CT） or basal diet supplemented with either whole full fat soybean （WFS）, full fat expanded soybean （FPS） or whole full fat soybean with whole cottonseed and full fat expanded soybean （MIX）. The composition of the milk fat was analyzed by gas chromatography. Relative to control, the conjugated linoleic acid （CLA） concentration in milk fat from cows on FPS was significantly increased by 83.88% （P〈 0.05）. The proportions of C12：0 were decreased by 35.7, 35.51, and 38.65% in milk fat from cows on WFS, MIX, and FPS compared with cows on CT. Similar decreases in C 14：0 were 23.83, 24.85, and 31.48% in WFS, MIX, and FPS treatments, respectively. Feeding vegetable oilseeds increased the proportion of healthy FAs （mainly CLA）, whereas decreased the concentratiofi of C12：0 and C14：0. Therefore, milk and dairy products would have higher nutritive and therapeutic value.展开更多
Protein phosphorylation is an important post-translational modification that regulates milk protein structure and function.The objective of this study was to analyze the presence of phosphorylated casein.Bovine milk p...Protein phosphorylation is an important post-translational modification that regulates milk protein structure and function.The objective of this study was to analyze the presence of phosphorylated casein.Bovine milk proteins were first separated by SDS polyacrylamide gel electrophoresis.After in gels digestion and extraction,phosphorylated peptides were enriched by titanium dioxide and identified by ultra performance liquid chromatography coupled with nano electrospray ionization tandem mass spectrometry.This method ensured the identification of 20 phosphorylated peptides,including 7 phosphorylated forms of α_s1-casein,8 α_s2-casein,and 5 β-casein.Eight phosphorylated sites derived from 3 α_s1-caseins,3 α_s2-caseins,and 2 β-caseins were also identified,and localized on residues Ser^61,Ser^63 and Ser^130 in α_s1-casein;Thr^145,Ser^146 and Ser^158 in α_s2-casein;and Ser^50 and Thr^56 in β-casein.These findings provide valuable information for investigating casein phosphorylation of the bovine milk.展开更多
基金supported by the National Natural Science Foundation of China(31261140365)the National Basic Research Program of China(2011CB100804)the Agricultural Science and Technology Innovation Program of Chinese Academy of Agricultural Sciences(ASTIP-IAS12)
文摘The partial 16 S r RNA gene sequences（100 to 500 bp） were widely used to reveal rumen bacterial composition influenced by diets, while quantification of the changed uncultured bacteria was inconvenient due to difficult designing of specific primers based on short sequences. This study evaluated the effect of forage resources on rumen bacterial diversity and developed new strategy for primer design based on short sequences to quantify the changed uncultured bacteria. Denaturing gradient gel electrophoresis（DGGE） analysis and subsequent band sequencing were used to reveal the distinct rumen bacteria composition in cows fed with two forage sources（single corn stover vs. mixed forages including alfalfa hay and corn silage）. The bacterial diversity in the rumen of dairy cows fed with corn stover was lower than that with mixed forages（P0.05）. The bacterium named R-UB affiliating to uncultured Succinivibrionaceae was identified, and it was abundant in the rumen of cows fed with mixed forages compared to corn stover. The full length 16 S r RNA gene sequences with identity of 97% to the R-UB 16 S r RNA gene sequence were obtained from Gen Bank and used to design specific primers to quantify uncultured bacterium R-UB. All sequences of amplicon from the new primers were of 100% identity to R-UB sequences indicating the high specificity of new primers. Quantitative PCR confirmed that abundance of R-UB in the rumen of cows fed with corn stover was lower than those fed with mixed forages（P0.01）. New strategy for designing primers based on partial 16 S r RNA genes to quantify targeted uncultured bacteria was successfully developed. The rumen bacteria descending significantly in the cows fed corn stover compared to those fed mixed forages was identified as uncultured R-UB from Succinivibrionaceae.
基金supported by the National Key Technology R&D Program of China (2012BAD12B02-5)the Research Program of the State Key Laboratory of Animal Nutrition, China (2004DA125184G1103)the Synergetic Innovation Center of Food Safety and Nutrition, China
文摘Dairy cows undergo tremendous changes in physiological, metabolism and the immune function from pregnancy to lac- tation that are associated with cows being susceptible to metabolic and infectious diseases. The objective of this study is to investigate the changes of plasma proteome on 21 d before expected calving and 1 d after calving from dairy cows using an integrated proteomic approach consisting of minor abundance protein enrichment by ProteoMiner beads, protein labeling by isobaric tags for relative and absolute quantification, and protein identification by liquid chromatography coupled with tandem mass spectrometry. Nineteen proteins were changed around the time of calving. These proteins were asso- ciated with response to stress, including acute-phase response and defense response, based on the proteins annotation. In particular, three up-regulated proteins after calving including factor V, a2-antiplasmin and prothrombin were assigned into the complement and coagulation pathway. These results may provide new information in elucidating host response to lactation and parturition stress, and inflammatory-like conditions at the protein level. Differential proteins may serve as potential markers to regulate the lactation and parturition stress in periparturient dairy cows.
基金grants from Ministry of Sciences and Technology, China (2006BAD12B08, 2006BAD04A03)Ministry of Agriculture, China (Nyhzx07-036)
文摘A PCR method for detection of bovis, sheep, pig, and chicken derived materials in feedstuff was established, and the existing method was improved according to the research on general primer and species-specific primers. First, general primer designed according to 16S rRNA gene sequence of hovis, sheep, pig, chicken, fish, and horse mtDNA was used for primary detection of animal derived materials in feedstuff. Species-specific primers designed according to conserved sequence of mtDNA of bovis, sheep, pig, and chicken were used for amplification of a 271,274, 149, and 266 bp fragment, respectively. Further confirmation of the detection result was then carried out. PCR method for detection of animal derived materials in unknown feedstuff was developed by using general primer, relevant PCR system, and PCR condition. Also a PCR method for detection of each species （bovines, sheep, pig, and chicken） was designed by using our speciesspecific primers. High sensitivity and specificity of our method were confirmed with a minimum detection level of 0.1%. Method for detection of animal derived materials in this research is not only cheap and easy for operation but also precise and reliable results can be obtained. It could be one of the effective methods for the detection of animal derived materials in feedstuff.
文摘To determine the effect of dietary supplementation with vegetable oilseeds on the composition of bovine milk fatty acids （FAs）, 40 Holstein dairy cows were used with a complete randomized design. At the beginning of the experiment, the cows were 150±25 day in milk （DIM）. Total duration of the experiment was six weeks. Measurements were made during the last three weeks. Cows in four treatments were fed with a basal diet （CT） or basal diet supplemented with either whole full fat soybean （WFS）, full fat expanded soybean （FPS） or whole full fat soybean with whole cottonseed and full fat expanded soybean （MIX）. The composition of the milk fat was analyzed by gas chromatography. Relative to control, the conjugated linoleic acid （CLA） concentration in milk fat from cows on FPS was significantly increased by 83.88% （P〈 0.05）. The proportions of C12：0 were decreased by 35.7, 35.51, and 38.65% in milk fat from cows on WFS, MIX, and FPS compared with cows on CT. Similar decreases in C 14：0 were 23.83, 24.85, and 31.48% in WFS, MIX, and FPS treatments, respectively. Feeding vegetable oilseeds increased the proportion of healthy FAs （mainly CLA）, whereas decreased the concentratiofi of C12：0 and C14：0. Therefore, milk and dairy products would have higher nutritive and therapeutic value.
基金supported by an earmark fund for the National Key Basic Research Program of China(2011CB100805)
文摘Protein phosphorylation is an important post-translational modification that regulates milk protein structure and function.The objective of this study was to analyze the presence of phosphorylated casein.Bovine milk proteins were first separated by SDS polyacrylamide gel electrophoresis.After in gels digestion and extraction,phosphorylated peptides were enriched by titanium dioxide and identified by ultra performance liquid chromatography coupled with nano electrospray ionization tandem mass spectrometry.This method ensured the identification of 20 phosphorylated peptides,including 7 phosphorylated forms of α_s1-casein,8 α_s2-casein,and 5 β-casein.Eight phosphorylated sites derived from 3 α_s1-caseins,3 α_s2-caseins,and 2 β-caseins were also identified,and localized on residues Ser^61,Ser^63 and Ser^130 in α_s1-casein;Thr^145,Ser^146 and Ser^158 in α_s2-casein;and Ser^50 and Thr^56 in β-casein.These findings provide valuable information for investigating casein phosphorylation of the bovine milk.
