目的 利用巴斯德毕赤 (Pichia pastoris)酵母真核表达系统 ,构建真核表达载体 p PIC9K与人溶菌酶基因 (humanlysozyme,h L Y)的重组质粒 p PIC9K- h L Y,表达出具有活性的人溶菌酶 .方法 此项研究在本实验室构建的人溶菌酶(h L Y)基因...目的 利用巴斯德毕赤 (Pichia pastoris)酵母真核表达系统 ,构建真核表达载体 p PIC9K与人溶菌酶基因 (humanlysozyme,h L Y)的重组质粒 p PIC9K- h L Y,表达出具有活性的人溶菌酶 .方法 此项研究在本实验室构建的人溶菌酶(h L Y)基因与 p UC19的重组质粒 p UC19- h L Y的基础上 ,通过设计一对引物 ,用多聚酶链式反应 (PCR)扩增出人溶菌酶全基因片段后 ,将其克隆到巴斯德毕赤酵母分泌型表达载体p PIC9K中 ,构建重组质粒 p PIC9K- h L Y,经 Bg1 酶切线性化后 ,用电穿孔法将其转入毕赤酵母细胞内 .通过 G418筛选 ,选到的高拷贝转化子经 PCR检测人溶菌基因与毕赤酵母染色体是否稳定整合 ,阳性克隆经甲醇诱导进行表达 .结果 序列测定 ,经 PCR扩增后的人溶菌酶基因与文献发表的序列相比 ,缺失 4个碱基 .我们决定采用重组 PCR法予以纠正 ,经序列测定结果正确 .用 PCR检测 ,人溶菌酶基因与毕赤酵母染色体稳定整合 .用甲醇诱导表达并以 SDS- PAGE分析 ,在 Mr为 15 0 0 0左右出现一条蛋白带 ,表达量约占上清总蛋白的 0 .47.Western Blot证实表达产物具有天然人溶菌酶的抗原性 .用黄色小球菌平板溶圈法鉴定表达产物具有人溶菌酶的活性 .结论 成功的构建了重组质粒 p PIC9K- h L展开更多
IM To study the distribution of arylsulfatase, βgalactosidase and lysozyme in gastric cancer cells, and its relationship to differentiation and invasion of gastric cancer cells.METHODS Histochemical, immunohistoc...IM To study the distribution of arylsulfatase, βgalactosidase and lysozyme in gastric cancer cells, and its relationship to differentiation and invasion of gastric cancer cells.METHODS Histochemical, immunohistochemical and ruthenium red (RR) electrocytochemical technique for three types of hydrolases and proteoglycans in pericancerous matrix in 33 cases of gastric cancer were observed under light and electron microscopy.RESULTS The expression intensities of arylsulfatase, βglactosidase and lysozyme in mucinous cell carcinomas were more intensive than those in welldifferentiated and poorlydifferentiated adenocarcinomas (P<005-001). The fibrous tissues smooth muscle and proteoglycans close to the cancer cells were degraded. They were found in the region far from the cancer cells. Expression of three enzymes mentioned above was low in adenocarcinoma cells, and fibrous tissues and RR granules were present and intact near the welldifferentiated and poorly differentiated adenocarcinoma cells.CONCLUSION Mucinous cell carcinoma may release various hydrolases into extracellular matrix, inducing degradation of pericancerous matrix and facilitating cancer cell invasion and metastasis..展开更多
Background Optimal gut health is important to maximize growth performance and feed efficiency in broiler chickens.A total of 1,365 one-day-old male Ross 308 broiler chickens were randomly divided into 5 treatments gro...Background Optimal gut health is important to maximize growth performance and feed efficiency in broiler chickens.A total of 1,365 one-day-old male Ross 308 broiler chickens were randomly divided into 5 treatments groups with 21 replicates,13 birds per replicate.The present research investigated effects of microbial muramidase or a precision glycan alone or in combination on growth performance,apparent total tract digestibility,total blood carotenoid content,intestinal villus length,meat quality and gut microbiota in broiler chickens.Treatments included:NC:negative control(basal diet group);PC:positive control(basal diet+0.02%probiotics);MR:basal diet+0.035%microbial muramidase;PG:basal diet+0.1%precision glycan;and MRPG:basal diet+0.025%MR+0.1%PG,respectively.Results MRPG group increased the body weight gain and feed intake(P<0.05)compared with NC group.Moreover,it significantly increased total serum carotenoid(P<0.05)and MRPG altered the microbial diversity in ileum contents.The MRPG treatment group increased the abundance of the phylum Firmicutes,and family Lachnospiraceae,Ruminococcaceae,Oscillospiraceae,Lactobacillaceae,Peptostreptococcaceae and decreased the abundance of the phylum Campilobacterota,Bacteroidota and family Bacteroidaceae.Compared with the NC group,the chickens fed MRPG showed significantly increased in duodenum villus length at end the trial.Conclusion In this study,overall results showed that the synergetic effects of MR and PG showed enhancing growth performance,total serum carotenoid level and altering gut microbiota composition of broilers.The current research indicates that co-supplementation of MR and PG in broiler diets enhances intestinal health,consequently leading to an increased broiler production.展开更多
Microorganisms play an important role in the conversion of grape juice into wine. Different species of yeast are mainly responsible for the production of ethanol. Lactic acid bacteria also occur regularly in must or w...Microorganisms play an important role in the conversion of grape juice into wine. Different species of yeast are mainly responsible for the production of ethanol. Lactic acid bacteria also occur regularly in must or wine. They are mostly undesirable due to their capacity to produce wine-spoiling compounds. Especially strains of Lactobacillus brevis are able to produce biogenic amines as well as precursors of ethyl carbamate and different off-flavours (N-heterocycles, volatile phenols). By excessive formation of acetic acid some lactobacilli may even induce slow/stuck grape juice fermentations. In conventional winemaking, additions of sulphite or lysozyme are used to inhibit the growth of spoilage microorganisms. There is a strong interest in finding alternatives, because of the reduced activity of lysozyme in the wine milieu, a limited growth-inhibiting activity against lactic acid bacteria, and some health risks described regarding the application of sulphite. We found that a culture supernatant of Streptomyces albidoflavus B 578 lysed all bacteria previously isolated from must and wine samples (Acetobacter sp., Lactobacillus sp., Leuconostoc sp., Oenococcus oeni, Pediococcus sp., Staphylococcus sp.) including 35 strains of L. brevis. Two bacteriolytic exoenzymes were isolated and characterized from the streptomycete: a muramidase (24 kDa) and a protease (17 kDa). Both hydrolyzed cell wall components of L. brevis (peptidoglycan, S-layer proteins) and were active under wine-relevant conditions.展开更多
文摘目的 利用巴斯德毕赤 (Pichia pastoris)酵母真核表达系统 ,构建真核表达载体 p PIC9K与人溶菌酶基因 (humanlysozyme,h L Y)的重组质粒 p PIC9K- h L Y,表达出具有活性的人溶菌酶 .方法 此项研究在本实验室构建的人溶菌酶(h L Y)基因与 p UC19的重组质粒 p UC19- h L Y的基础上 ,通过设计一对引物 ,用多聚酶链式反应 (PCR)扩增出人溶菌酶全基因片段后 ,将其克隆到巴斯德毕赤酵母分泌型表达载体p PIC9K中 ,构建重组质粒 p PIC9K- h L Y,经 Bg1 酶切线性化后 ,用电穿孔法将其转入毕赤酵母细胞内 .通过 G418筛选 ,选到的高拷贝转化子经 PCR检测人溶菌基因与毕赤酵母染色体是否稳定整合 ,阳性克隆经甲醇诱导进行表达 .结果 序列测定 ,经 PCR扩增后的人溶菌酶基因与文献发表的序列相比 ,缺失 4个碱基 .我们决定采用重组 PCR法予以纠正 ,经序列测定结果正确 .用 PCR检测 ,人溶菌酶基因与毕赤酵母染色体稳定整合 .用甲醇诱导表达并以 SDS- PAGE分析 ,在 Mr为 15 0 0 0左右出现一条蛋白带 ,表达量约占上清总蛋白的 0 .47.Western Blot证实表达产物具有天然人溶菌酶的抗原性 .用黄色小球菌平板溶圈法鉴定表达产物具有人溶菌酶的活性 .结论 成功的构建了重组质粒 p PIC9K- h L
文摘IM To study the distribution of arylsulfatase, βgalactosidase and lysozyme in gastric cancer cells, and its relationship to differentiation and invasion of gastric cancer cells.METHODS Histochemical, immunohistochemical and ruthenium red (RR) electrocytochemical technique for three types of hydrolases and proteoglycans in pericancerous matrix in 33 cases of gastric cancer were observed under light and electron microscopy.RESULTS The expression intensities of arylsulfatase, βglactosidase and lysozyme in mucinous cell carcinomas were more intensive than those in welldifferentiated and poorlydifferentiated adenocarcinomas (P<005-001). The fibrous tissues smooth muscle and proteoglycans close to the cancer cells were degraded. They were found in the region far from the cancer cells. Expression of three enzymes mentioned above was low in adenocarcinoma cells, and fibrous tissues and RR granules were present and intact near the welldifferentiated and poorly differentiated adenocarcinoma cells.CONCLUSION Mucinous cell carcinoma may release various hydrolases into extracellular matrix, inducing degradation of pericancerous matrix and facilitating cancer cell invasion and metastasis..
基金supported by Basic Science Research Program through the National Research Foundation of Korea(NRF)funded by the Ministry of Education(NRF-RS-2023-00275307)。
文摘Background Optimal gut health is important to maximize growth performance and feed efficiency in broiler chickens.A total of 1,365 one-day-old male Ross 308 broiler chickens were randomly divided into 5 treatments groups with 21 replicates,13 birds per replicate.The present research investigated effects of microbial muramidase or a precision glycan alone or in combination on growth performance,apparent total tract digestibility,total blood carotenoid content,intestinal villus length,meat quality and gut microbiota in broiler chickens.Treatments included:NC:negative control(basal diet group);PC:positive control(basal diet+0.02%probiotics);MR:basal diet+0.035%microbial muramidase;PG:basal diet+0.1%precision glycan;and MRPG:basal diet+0.025%MR+0.1%PG,respectively.Results MRPG group increased the body weight gain and feed intake(P<0.05)compared with NC group.Moreover,it significantly increased total serum carotenoid(P<0.05)and MRPG altered the microbial diversity in ileum contents.The MRPG treatment group increased the abundance of the phylum Firmicutes,and family Lachnospiraceae,Ruminococcaceae,Oscillospiraceae,Lactobacillaceae,Peptostreptococcaceae and decreased the abundance of the phylum Campilobacterota,Bacteroidota and family Bacteroidaceae.Compared with the NC group,the chickens fed MRPG showed significantly increased in duodenum villus length at end the trial.Conclusion In this study,overall results showed that the synergetic effects of MR and PG showed enhancing growth performance,total serum carotenoid level and altering gut microbiota composition of broilers.The current research indicates that co-supplementation of MR and PG in broiler diets enhances intestinal health,consequently leading to an increased broiler production.
基金support by the Deutsche Forschungsgemeinschaft(DFGKo 785/17-1)the FEI(Forschungskreis der Ernährungsindustrie E.V.Bonn)Projekt AIF-FV 15833N.
文摘Microorganisms play an important role in the conversion of grape juice into wine. Different species of yeast are mainly responsible for the production of ethanol. Lactic acid bacteria also occur regularly in must or wine. They are mostly undesirable due to their capacity to produce wine-spoiling compounds. Especially strains of Lactobacillus brevis are able to produce biogenic amines as well as precursors of ethyl carbamate and different off-flavours (N-heterocycles, volatile phenols). By excessive formation of acetic acid some lactobacilli may even induce slow/stuck grape juice fermentations. In conventional winemaking, additions of sulphite or lysozyme are used to inhibit the growth of spoilage microorganisms. There is a strong interest in finding alternatives, because of the reduced activity of lysozyme in the wine milieu, a limited growth-inhibiting activity against lactic acid bacteria, and some health risks described regarding the application of sulphite. We found that a culture supernatant of Streptomyces albidoflavus B 578 lysed all bacteria previously isolated from must and wine samples (Acetobacter sp., Lactobacillus sp., Leuconostoc sp., Oenococcus oeni, Pediococcus sp., Staphylococcus sp.) including 35 strains of L. brevis. Two bacteriolytic exoenzymes were isolated and characterized from the streptomycete: a muramidase (24 kDa) and a protease (17 kDa). Both hydrolyzed cell wall components of L. brevis (peptidoglycan, S-layer proteins) and were active under wine-relevant conditions.
