The gene encoding the 18 kDa protein of Taenia solium metacestodes was amplified by RT-PCR and cloned into the pGEM-T vector for sequencing.The recombinant plasmid named pGEX-CE18 was constructed and transformed into ...The gene encoding the 18 kDa protein of Taenia solium metacestodes was amplified by RT-PCR and cloned into the pGEM-T vector for sequencing.The recombinant plasmid named pGEX-CE18 was constructed and transformed into E.coli BL21 for in vitro expression.SDS-PAGE and Western blot were employed for analyzing the recombinant protein,which was then used for development of an indirect ELISA for detection of anti-cysticercosis antibodies.The results showed that the recombinant protein of interest was 35 kDa in size,accounting for 28%of total bacteria proteins,and reacted with positive sera against cysticercosis.Using the newly-constructed indirect ELISA and a commercially available ELISA kit,paired analyses of 178 serum samples indicated that the concordant rate was 98.83%and the ELISA exhibited good specificity and sensitivity,supporting its utility and application for diagnosis of cysticercosis.展开更多
OBJECTIVE Metrnl is a novel secreted protein with limited researches.In this study,we investigated metrnl tissue expression pattern in humans,and exploredthe possible role of its highest expression using animal models...OBJECTIVE Metrnl is a novel secreted protein with limited researches.In this study,we investigated metrnl tissue expression pattern in humans,and exploredthe possible role of its highest expression using animal models.METHODS We examined metrnl tissue expression pattern in a human tissue microarray containing 19types of tissues from 69 donors,and verified the highest expression in fresh human and mouse tissues.We then created an animal model of cell-specific knockout mice to study the role of metrnl.RESULTS Metrnl was the highest expressed in human gastrointestinal tract,and specifical y expressed in the intestinal epithelium.Consistently,Metrnl expression was also the highest expressed in mouse gastrointestinal tract among the detected tissues of 14 types.We developed intestinal epithelial cellspecific metrnl knockout mice with Vil in-Cre.In this animal model,metrnl levels displayed a statistically significant reduction in gut fluid,but not in blood serum.This cell specific deletion of metrnl did not affect body weight,food intake,blood glucose,colon length and histology,intestinal permeability,mucus production and mucin 2 expression under physiological conditions,but markedly reduced the expression of antimicrobial peptides,such as regenerating islet-derived 3 gamma and lactotransferrin.CONCLUSION Metrnl is rich in intestinal epithelial cells of humans and mice,mainly contributing to local gut metrnl level,and less affecting systemic circulating metrnl level.Metrnl plays a role in maintaining gut antimicrobial peptides.展开更多
基于巴斯德毕赤酵母表达系统探讨了屋尘螨过敏原Der p 1的重组表达与纯化。首先,采用毕赤酵母GS115表达密码子优化的全长编码基因PreProDer p 1,其产量可达100 mg/L,进一步共表达分子伴侣实现产量提高到140 mg/L,并实现3 L反应器发酵产...基于巴斯德毕赤酵母表达系统探讨了屋尘螨过敏原Der p 1的重组表达与纯化。首先,采用毕赤酵母GS115表达密码子优化的全长编码基因PreProDer p 1,其产量可达100 mg/L,进一步共表达分子伴侣实现产量提高到140 mg/L,并实现3 L反应器发酵产量提高到1 g/L。其次,对上述重组蛋白发酵液分别使用阳离子交换层析及亲和层析进行了纯化工艺优化,目的蛋白得率达到60.7%。本研究为后续PreProDre p 1诊断试剂盆的开发提供了参考。展开更多
基金supported by National High-tech Research and Development Plan(863 Project)(2006AA10A207)
文摘The gene encoding the 18 kDa protein of Taenia solium metacestodes was amplified by RT-PCR and cloned into the pGEM-T vector for sequencing.The recombinant plasmid named pGEX-CE18 was constructed and transformed into E.coli BL21 for in vitro expression.SDS-PAGE and Western blot were employed for analyzing the recombinant protein,which was then used for development of an indirect ELISA for detection of anti-cysticercosis antibodies.The results showed that the recombinant protein of interest was 35 kDa in size,accounting for 28%of total bacteria proteins,and reacted with positive sera against cysticercosis.Using the newly-constructed indirect ELISA and a commercially available ELISA kit,paired analyses of 178 serum samples indicated that the concordant rate was 98.83%and the ELISA exhibited good specificity and sensitivity,supporting its utility and application for diagnosis of cysticercosis.
基金The project supported by National Natural Science Foundation of China(81130061,81202572,81373414)
文摘OBJECTIVE Metrnl is a novel secreted protein with limited researches.In this study,we investigated metrnl tissue expression pattern in humans,and exploredthe possible role of its highest expression using animal models.METHODS We examined metrnl tissue expression pattern in a human tissue microarray containing 19types of tissues from 69 donors,and verified the highest expression in fresh human and mouse tissues.We then created an animal model of cell-specific knockout mice to study the role of metrnl.RESULTS Metrnl was the highest expressed in human gastrointestinal tract,and specifical y expressed in the intestinal epithelium.Consistently,Metrnl expression was also the highest expressed in mouse gastrointestinal tract among the detected tissues of 14 types.We developed intestinal epithelial cellspecific metrnl knockout mice with Vil in-Cre.In this animal model,metrnl levels displayed a statistically significant reduction in gut fluid,but not in blood serum.This cell specific deletion of metrnl did not affect body weight,food intake,blood glucose,colon length and histology,intestinal permeability,mucus production and mucin 2 expression under physiological conditions,but markedly reduced the expression of antimicrobial peptides,such as regenerating islet-derived 3 gamma and lactotransferrin.CONCLUSION Metrnl is rich in intestinal epithelial cells of humans and mice,mainly contributing to local gut metrnl level,and less affecting systemic circulating metrnl level.Metrnl plays a role in maintaining gut antimicrobial peptides.
文摘基于巴斯德毕赤酵母表达系统探讨了屋尘螨过敏原Der p 1的重组表达与纯化。首先,采用毕赤酵母GS115表达密码子优化的全长编码基因PreProDer p 1,其产量可达100 mg/L,进一步共表达分子伴侣实现产量提高到140 mg/L,并实现3 L反应器发酵产量提高到1 g/L。其次,对上述重组蛋白发酵液分别使用阳离子交换层析及亲和层析进行了纯化工艺优化,目的蛋白得率达到60.7%。本研究为后续PreProDre p 1诊断试剂盆的开发提供了参考。
