The paper was to obtain the VP1 protein of FMDV serotype A with high activity. With recombinant plasmid pMD19A-T-vp1 as the tem- plate, vpl gene fragment amplified by PCR was connected into prokaryotic expression vect...The paper was to obtain the VP1 protein of FMDV serotype A with high activity. With recombinant plasmid pMD19A-T-vp1 as the tem- plate, vpl gene fragment amplified by PCR was connected into prokaryotic expression vector pET28a to construct recombinant plasmid pET-A-vpl. The E. coli BL21 （DE3） strain containing recombinant plasmid pET-A-vpl were induced by IPTG. SDS-PAGE showed that VP1 protein was ex- pressed in the form of inclusion body, and its molecular weight was about 29 ku. Based on the optimizing IPTG concentration and expression time, the largest expression of VP1 protein was induced by 0.3 mmol/L IPTG for 6 h at 37 ℃. Western-Blot analysis indicated that the expression of VP1 protein could be specifically recognized by positive serum of FMDV serotype A. ELISA test showed that VP1 inclusion body protein had high activity after purification by washing and renaturation by urea concentration gradient dialysis.展开更多
[Objective] The research aimed to induce the expression of FMDV structural protein VP1 in E.coli and purify the protein,then detect the activity.[Method] The fragment coding VP1 was amplified by PCR and doubly digeste...[Objective] The research aimed to induce the expression of FMDV structural protein VP1 in E.coli and purify the protein,then detect the activity.[Method] The fragment coding VP1 was amplified by PCR and doubly digested with BamH Ⅰ and XhoⅠ,then cloned into expression vector pGEX-4T-1 and pPROExHTb respectively to get recombinant plasmid pGEX-4T-1-VP1 and pPROExHTb-VP1.The recombinant plasmid pGEX-4T-1-VP1 and pPROExHTb-VP1 was transformed into E.coli BL21(DE3)and induced by IPTG,fusion protein was identifie...展开更多
Heat stress transcription factors (Hsfs) are the central regulators of defense response to heat stress. We identified a total of 25 rice Hsf genes by genome-wide analysis of rice (Oryza sativa L.) genome, including th...Heat stress transcription factors (Hsfs) are the central regulators of defense response to heat stress. We identified a total of 25 rice Hsf genes by genome-wide analysis of rice (Oryza sativa L.) genome, including the subspecies of O. japonica and O. indica. Proteins encoded by OsHsfs were divided into three classes according to their structures. Digital Northern analysis showed that OsHsfs were expressed constitutively. The expressions of these OsHsfs in response to heat stress and oxidative stress differed among the members of the gene family. Promoter analysis identified a number of stress-related cis-elements in the promoter regions of these OsHsfs. No significant correlation, however, was found between the heat-shock responses of genes and their cis-elements. Overall, our results provide a foundation for future research of OsHsfs function.展开更多
Parotid secretory protein (PSP) secreted abundantly in saliva, whose function is related with the anti-bacterial effect. The PSP cDNA has been isolated from pig parotid glands by 3′ and 5′ rapid amplification of cDN...Parotid secretory protein (PSP) secreted abundantly in saliva, whose function is related with the anti-bacterial effect. The PSP cDNA has been isolated from pig parotid glands by 3′ and 5′ rapid amplification of cDNA end (RACE), based on the conserved signal peptide region among the known mammalian PSP. The result of homologous comparison shows that pig PSP and human PSP shares the high identity at the level of the primary, secondary and tertiary protein structure. A search for functionally significant protein motifs revealed a unique amino acid sequence pattern consisting of the residues Leu-X(6)-Leu-X(6)-Leu- X(7)-Leu-X(6)-Leu-X(6)-Leu near the amino-terminal portion of the protein, which is important to its function. RT-PCR, Dot blot and Northern blot analysis demonstrated that PSP was strongly expressed in parotid glands, but not in other tissues.展开更多
目的:以纯化的壶瓶碎米荠含硒蛋白(selenium-containing protein from Cardamine hupingshanensis,S P C H)为研究对象,对其结构特性和缓解运动性疲劳作用进行评价。方法:采用聚丙烯酰胺凝胶电泳(polyacrylamide gelelectrophoresis,PA...目的:以纯化的壶瓶碎米荠含硒蛋白(selenium-containing protein from Cardamine hupingshanensis,S P C H)为研究对象,对其结构特性和缓解运动性疲劳作用进行评价。方法:采用聚丙烯酰胺凝胶电泳(polyacrylamide gelelectrophoresis,PAGE)和十二烷基硫酸钠-PAGE(sodium dodecyl sulfate-PAGE,SDS-PAGE)、氨基酸组成分析、基质辅助激光解吸电离飞行时间质谱(matrix-assisted laser desorption/ionization time of flight mass spectrometry,MALDI-TOF-MS)分析,对SPCH的纯度、亚基组成、氨基酸组成及含量进行评价并初步预测SPCH的匹配蛋白。采用小鼠负重游泳实验,通过检测游泳时间、血乳酸(blood lactic acid,BLA)、肝糖原和血尿素氮(blood urea nitrogen,BUN)水平,评价SPCH对小鼠运动性疲劳的影响。结果:SPCH可能由3个分子质量分别为37、39、40 k D的亚基组成,与之相匹配的蛋白可能是DING protein、Predicted protein和Chalcone synthase。SPCH能显著延长小鼠负重游泳时间(P<0.01),同时增强清除乳酸的能力(P<0.01),增加肝糖原含量(P<0.01),还具有降低BUN水平的能力(P<0.05)。结论:SPCH对小鼠具有较好的缓解运动性疲劳作用,可考虑将其开发成为缓解运动性疲劳的营养补充剂。展开更多
基金Supported by NSFC-Joint Personnel Training Fund of Henan Province(U1204327)Special Fund for Construction of Provincial Key Laboratory in Henan Province(122300413217)
文摘The paper was to obtain the VP1 protein of FMDV serotype A with high activity. With recombinant plasmid pMD19A-T-vp1 as the tem- plate, vpl gene fragment amplified by PCR was connected into prokaryotic expression vector pET28a to construct recombinant plasmid pET-A-vpl. The E. coli BL21 （DE3） strain containing recombinant plasmid pET-A-vpl were induced by IPTG. SDS-PAGE showed that VP1 protein was ex- pressed in the form of inclusion body, and its molecular weight was about 29 ku. Based on the optimizing IPTG concentration and expression time, the largest expression of VP1 protein was induced by 0.3 mmol/L IPTG for 6 h at 37 ℃. Western-Blot analysis indicated that the expression of VP1 protein could be specifically recognized by positive serum of FMDV serotype A. ELISA test showed that VP1 inclusion body protein had high activity after purification by washing and renaturation by urea concentration gradient dialysis.
基金Supported by National Supporting Plan(2006BAD06A14)Transgenic Major Projects(2008ZX08011-004)~~
文摘[Objective] The research aimed to induce the expression of FMDV structural protein VP1 in E.coli and purify the protein,then detect the activity.[Method] The fragment coding VP1 was amplified by PCR and doubly digested with BamH Ⅰ and XhoⅠ,then cloned into expression vector pGEX-4T-1 and pPROExHTb respectively to get recombinant plasmid pGEX-4T-1-VP1 and pPROExHTb-VP1.The recombinant plasmid pGEX-4T-1-VP1 and pPROExHTb-VP1 was transformed into E.coli BL21(DE3)and induced by IPTG,fusion protein was identifie...
基金Project (No. 30471118) supported by the National Natural Science Foundation of China
文摘Heat stress transcription factors (Hsfs) are the central regulators of defense response to heat stress. We identified a total of 25 rice Hsf genes by genome-wide analysis of rice (Oryza sativa L.) genome, including the subspecies of O. japonica and O. indica. Proteins encoded by OsHsfs were divided into three classes according to their structures. Digital Northern analysis showed that OsHsfs were expressed constitutively. The expressions of these OsHsfs in response to heat stress and oxidative stress differed among the members of the gene family. Promoter analysis identified a number of stress-related cis-elements in the promoter regions of these OsHsfs. No significant correlation, however, was found between the heat-shock responses of genes and their cis-elements. Overall, our results provide a foundation for future research of OsHsfs function.
基金supported by the National Major Basic Research Development Program(Grant No.G20000161)Beijing Natural Science Foundation(Grant No.5030001).
文摘Parotid secretory protein (PSP) secreted abundantly in saliva, whose function is related with the anti-bacterial effect. The PSP cDNA has been isolated from pig parotid glands by 3′ and 5′ rapid amplification of cDNA end (RACE), based on the conserved signal peptide region among the known mammalian PSP. The result of homologous comparison shows that pig PSP and human PSP shares the high identity at the level of the primary, secondary and tertiary protein structure. A search for functionally significant protein motifs revealed a unique amino acid sequence pattern consisting of the residues Leu-X(6)-Leu-X(6)-Leu- X(7)-Leu-X(6)-Leu-X(6)-Leu near the amino-terminal portion of the protein, which is important to its function. RT-PCR, Dot blot and Northern blot analysis demonstrated that PSP was strongly expressed in parotid glands, but not in other tissues.
文摘目的:以纯化的壶瓶碎米荠含硒蛋白(selenium-containing protein from Cardamine hupingshanensis,S P C H)为研究对象,对其结构特性和缓解运动性疲劳作用进行评价。方法:采用聚丙烯酰胺凝胶电泳(polyacrylamide gelelectrophoresis,PAGE)和十二烷基硫酸钠-PAGE(sodium dodecyl sulfate-PAGE,SDS-PAGE)、氨基酸组成分析、基质辅助激光解吸电离飞行时间质谱(matrix-assisted laser desorption/ionization time of flight mass spectrometry,MALDI-TOF-MS)分析,对SPCH的纯度、亚基组成、氨基酸组成及含量进行评价并初步预测SPCH的匹配蛋白。采用小鼠负重游泳实验,通过检测游泳时间、血乳酸(blood lactic acid,BLA)、肝糖原和血尿素氮(blood urea nitrogen,BUN)水平,评价SPCH对小鼠运动性疲劳的影响。结果:SPCH可能由3个分子质量分别为37、39、40 k D的亚基组成,与之相匹配的蛋白可能是DING protein、Predicted protein和Chalcone synthase。SPCH能显著延长小鼠负重游泳时间(P<0.01),同时增强清除乳酸的能力(P<0.01),增加肝糖原含量(P<0.01),还具有降低BUN水平的能力(P<0.05)。结论:SPCH对小鼠具有较好的缓解运动性疲劳作用,可考虑将其开发成为缓解运动性疲劳的营养补充剂。
