[Objective] This study aimed to construct DNA vaccine of foot-and-mouth disease （FMD）.[Method] Plasmid carriers plESZP1 and pUTK3CP1 with PRV were constructed for FMDV P1 gene expression.Mice were immunized,and thei...[Objective] This study aimed to construct DNA vaccine of foot-and-mouth disease （FMD）.[Method] Plasmid carriers plESZP1 and pUTK3CP1 with PRV were constructed for FMDV P1 gene expression.Mice were immunized,and their antibody level was detected.The two eukaryotic expression plasmids constructed were transfected into Vero cells.PCR,IFA and Westem-blot were carried out to detect the transcription and expression of the objective gene.Balb/C mice were intramuscularly inoculated with the DNA plasmid which expressed the target gene correctly,and the antibody level in mice was detected by the means of ELISA and serum neutralization （SN）.[Result] DNA plasmid carrying P1 gene which encodes FMDV capsid protein caused specific body fluid immunoreaction in mice,and the antibody level of anti-FMDV had no difference in the mice induced by the two recombinant plasmids.[Conclusion] This study lays a foundation for evaluating the genetically modified vaccine by immunizing animals with recombinant PRV containing the FMDV P1 gene and recombinant virus.展开更多
[Objective] The aim was to study the prokaryotic expression of P1 gene of foot-and-mouth disease virus（FMDV）type Asia 1and the preparation of its antiserum.[Method]The P1 gene of FMDV type Asia 1 was obtained by gen...[Objective] The aim was to study the prokaryotic expression of P1 gene of foot-and-mouth disease virus（FMDV）type Asia 1and the preparation of its antiserum.[Method]The P1 gene of FMDV type Asia 1 was obtained by gene cloning techniques,and then cloned into pET-32a（＋）plasmid;subsequently the recombinant plasmid was transformed into E.coli BL21（DE3）;after the IPTG induction and protein purification,SDS-PAGE analysis was carried out;the ultrasonic wave was use to lyse the cultivated recombinant strain,and after the isolation and purification,this fusion protein was utilized to immunize New Zealand rabbits so as to prepare P1 protein antiserum.[Result]The positive clones were obtained;SDS-PAGE result showed that the target band was appeared at 105 kD;Western blot analysis showed that the antisera could bind to the expressed P1 fusion protein specifically;the ELISA titer of the rabbit anti-FMDV-P1 sera was approximately 1∶5 120.[Conclusion]This study had provided foundations for FMDV serological diagnostic methods and genetically engineered vaccine.展开更多
Objective: To investigate the p21WAF1 /CIP1gene DNA sequence change and their relationship with the phenotype of human osteosarcoma. Methods: p21WAF1 /CIP1gene DNA of 36 osteosarcoma spec- ...Objective: To investigate the p21WAF1 /CIP1gene DNA sequence change and their relationship with the phenotype of human osteosarcoma. Methods: p21WAF1 /CIP1gene DNA of 36 osteosarcoma spec- imens was examined by using polymerase chain reaction-single strand conformation polymorphism (PCR- SSCP) method. The PCR products were sequenced directly. Results: In p21WAF1 /CIP1 gene exon3 of 36 cases of human osteosarcoma, the change of C→T in the p21WAF1 /CIP1gene CDNA sequence of position 609th occurred in 17 cases with the incidence being 44.4%. In 10 normal blood samples, DNA sequence analysis showed the change of C→T in the p21WAF1 /CIP1gene CDNA sequence of position 609th occurred in 8 cases with the incidence being 80%. Conclusion: The novel location of p21WAF1 /CIP1gene polymorphism of osteosarcoma, but not mutation was de?ned, and this location might provide the meaningful reference for the further research of p21WAF1/CIP1 gene.p2lWAF1/CIP1基因DNA序列分析及其与骨肉瘤表型的关系展开更多
Background Mycoplasma pneumoniae is a common pathogen that caused community-acquired pneumonia （CAP）. P1 protein served as major adhesion and immunodominant protein in Mycoplasma pneumoniae, but little about P1 gene...Background Mycoplasma pneumoniae is a common pathogen that caused community-acquired pneumonia （CAP）. P1 protein served as major adhesion and immunodominant protein in Mycoplasma pneumoniae, but little about P1 gene was learned and the relationship between P1 genotype and macrolide resistance has yet to be explored.展开更多
Objective To investigate the effect of trans-acting factor(s) on rat glutathione S-transferase P1 gene (rGSTP1) transcription regulation in tumor cells.Methods The binding of trans-acting factor(s) to two enhancers...Objective To investigate the effect of trans-acting factor(s) on rat glutathione S-transferase P1 gene (rGSTP1) transcription regulation in tumor cells.Methods The binding of trans-acting factor(s) to two enhancers of the rGSTP1 gene, glutathione S-transferase P enhancer Ⅰ (GPEI) and glutathione S-transferase P enhancer Ⅱ-1 (GPEⅡ-1), was identified by an electrophoretic mobility shift assay (EMSA). The molecular weight of trans-acting factor was measured in a UV cross-linking experiment. Results Trans-acting factor interacting with the core sequence of GPEI (cGPEI) were found in human cervical adenocarcinoma cell line (HeLa) and rat hepatoma cell line (CBRH7919). These proteins were not expressed in normal rat liver. Although specific binding proteins that bound to GPEⅡ-1 were detected in all three cell types, a 64 kDa binding protein that exists in HeLa and CBRH7919 cells was absent in normal rat liver. Conclusion cGPEI, GPEII specific binding proteins expressed in HeLa and CBRH7919 cells may play an important role in the high transcriptional level of the rGSTP1 gene in tumor cells.展开更多
Objective: To investigate the possible relationship between deletion of MTS/p16 gene and progression of endometrial carcinoma Methods: Forty six primary endometrial carcinoma, 7 tumor adjacent endometrial tissue,...Objective: To investigate the possible relationship between deletion of MTS/p16 gene and progression of endometrial carcinoma Methods: Forty six primary endometrial carcinoma, 7 tumor adjacent endometrial tissue, 10 normal endometrial tissue specimen and 5 xenografts from patients with endometrial carcinoma were examined for homozygous deletion of MTS/p16 gene by polymerase chain reaction based analysis Results: Of 46 endometrial cancer specimens, 9 showed homozygous deletion, no deletion was detected in the tumor adjacent and normal endometial tissues Nor was it detected in well differentiated endometrial carcinoma and all xenografts Conclusions: Deletion of MTS1/p16 gene might contribute to the progression of endometrial carcinoma and could be served as indicator for predicting prognosis展开更多
Polycyclic aromatic hydrocarbons(PAHs),dimethyl beazanthracene(DMBA)and benzo-(a)-pyrene(BaP),stimulate the expression of the P1-450 gene in human lung tumor cells(ChaGo)in culture.A concentration-and time-dependent i...Polycyclic aromatic hydrocarbons(PAHs),dimethyl beazanthracene(DMBA)and benzo-(a)-pyrene(BaP),stimulate the expression of the P1-450 gene in human lung tumor cells(ChaGo)in culture.A concentration-and time-dependent increase in the level of P1-450 specific mRNA sequences hasbeen observed in ChaGo cells treated with sublethal concentrations of DMBA.Results presentedsuggest that the parent compound causes the induction of P1-450 gene expression.The methylationpattern of the internal“-C-” residues of the “-CCGG-” sequence has been studied in a stretch ofabout 8000 base pairs(bp)of DNA sequence in and around the P1-450 gene in the control andDMBA-treated ChaGo cells.A comparative Southern blot analysis of the Msp Ⅰ/Hpa Ⅱ digestedDNA of the control and DMBA-treated ChaGo cells followed by hybridization with ^(32)P-labelledregion-specific probes,revealed that i)the internal “-C-” residues of the “-CCGG-” sequences ofdifferent regions of P1-450 gene were methylated to different degrees;ii)DMBA treatment of thecells induced hypomethylation of only the 5' end “-CCGG-” sequences of the gene;and iii)themethylation patterns of “-CCGG-” sequences of most of the coding region and of the 3' end regionof P1-450 gene were not affected by such DMBA treatment.展开更多
Objective: To study the relationship between the expression of p27Kip1 and p53, and the infiltration, metastasis and prognosis in gastric carcinoma. Methods: The expression of p27Kip1 and p53 at protein level was dete...Objective: To study the relationship between the expression of p27Kip1 and p53, and the infiltration, metastasis and prognosis in gastric carcinoma. Methods: The expression of p27Kip1 and p53 at protein level was determined by immunohistochemical assay (two-step method) in 100 cases of gastric carcinoma. Results: Of the 100 cases, the positive rate of p27Kip1 and p53 expressions were 44% and 49%, respectively. In the group of gastric carcinomas with deep infiltration (infiltration group), lymph nodes metastasis group (metastasis group) and death-within-5-years group (death group), the expression of p27Kip1 was statistically lower (_P<0.05). In the metastasis group and death group, the expression of p53 was significantly higher (P<0.05). The results of the monovariate analysis revealed that the 5-year survival rate of the high p27Kip1 expression group was 70.59%, which is higher than those of the low p27Kip1 expression group (54.55%) and the negative experession group (26%). The 5-year survival rate of the high p53 expression group was 19.23%, which was lower than those of the p53 low expression group (43.75%) and the negative group (53.19%). Cox multivariate analysis showed that p27Kip1, like p53, was an independent prognostic index. But p27Klpl protein expression was a stronger independent survival predictor (RR=3.06) than p53 expression (RR=2.33). Conclusion: The low expression of p27Kip1 and the high expression of p53 reflected the more frequent invasion and metastasis, which resulted in the reduced survival of patients. As an independent markers of the gastric carcinoma, the expression of p27Kip1 is more useful than that of p53 in the prognosis prediction of gastric carcinoma.展开更多
文摘[Objective] This study aimed to construct DNA vaccine of foot-and-mouth disease （FMD）.[Method] Plasmid carriers plESZP1 and pUTK3CP1 with PRV were constructed for FMDV P1 gene expression.Mice were immunized,and their antibody level was detected.The two eukaryotic expression plasmids constructed were transfected into Vero cells.PCR,IFA and Westem-blot were carried out to detect the transcription and expression of the objective gene.Balb/C mice were intramuscularly inoculated with the DNA plasmid which expressed the target gene correctly,and the antibody level in mice was detected by the means of ELISA and serum neutralization （SN）.[Result] DNA plasmid carrying P1 gene which encodes FMDV capsid protein caused specific body fluid immunoreaction in mice,and the antibody level of anti-FMDV had no difference in the mice induced by the two recombinant plasmids.[Conclusion] This study lays a foundation for evaluating the genetically modified vaccine by immunizing animals with recombinant PRV containing the FMDV P1 gene and recombinant virus.
基金Supported by National Transgenic Major Program of China(2009ZX08007-006B)the National Natural Science Foundation of China(31072160)+2 种基金Science and Technique Foundation of Shandong Province(2009GG20002032)Natural Science Foundation ofShandong Province(Y2008D20)an Open Issue of State Key Laboratory of Veterinary Biotechnology Fund(SKLVBF200806)~~
文摘[Objective] The aim was to study the prokaryotic expression of P1 gene of foot-and-mouth disease virus（FMDV）type Asia 1and the preparation of its antiserum.[Method]The P1 gene of FMDV type Asia 1 was obtained by gene cloning techniques,and then cloned into pET-32a（＋）plasmid;subsequently the recombinant plasmid was transformed into E.coli BL21（DE3）;after the IPTG induction and protein purification,SDS-PAGE analysis was carried out;the ultrasonic wave was use to lyse the cultivated recombinant strain,and after the isolation and purification,this fusion protein was utilized to immunize New Zealand rabbits so as to prepare P1 protein antiserum.[Result]The positive clones were obtained;SDS-PAGE result showed that the target band was appeared at 105 kD;Western blot analysis showed that the antisera could bind to the expressed P1 fusion protein specifically;the ELISA titer of the rabbit anti-FMDV-P1 sera was approximately 1∶5 120.[Conclusion]This study had provided foundations for FMDV serological diagnostic methods and genetically engineered vaccine.
文摘Objective: To investigate the p21WAF1 /CIP1gene DNA sequence change and their relationship with the phenotype of human osteosarcoma. Methods: p21WAF1 /CIP1gene DNA of 36 osteosarcoma spec- imens was examined by using polymerase chain reaction-single strand conformation polymorphism (PCR- SSCP) method. The PCR products were sequenced directly. Results: In p21WAF1 /CIP1 gene exon3 of 36 cases of human osteosarcoma, the change of C→T in the p21WAF1 /CIP1gene CDNA sequence of position 609th occurred in 17 cases with the incidence being 44.4%. In 10 normal blood samples, DNA sequence analysis showed the change of C→T in the p21WAF1 /CIP1gene CDNA sequence of position 609th occurred in 8 cases with the incidence being 80%. Conclusion: The novel location of p21WAF1 /CIP1gene polymorphism of osteosarcoma, but not mutation was de?ned, and this location might provide the meaningful reference for the further research of p21WAF1/CIP1 gene.p2lWAF1/CIP1基因DNA序列分析及其与骨肉瘤表型的关系
文摘Background Mycoplasma pneumoniae is a common pathogen that caused community-acquired pneumonia （CAP）. P1 protein served as major adhesion and immunodominant protein in Mycoplasma pneumoniae, but little about P1 gene was learned and the relationship between P1 genotype and macrolide resistance has yet to be explored.
文摘Objective To investigate the effect of trans-acting factor(s) on rat glutathione S-transferase P1 gene (rGSTP1) transcription regulation in tumor cells.Methods The binding of trans-acting factor(s) to two enhancers of the rGSTP1 gene, glutathione S-transferase P enhancer Ⅰ (GPEI) and glutathione S-transferase P enhancer Ⅱ-1 (GPEⅡ-1), was identified by an electrophoretic mobility shift assay (EMSA). The molecular weight of trans-acting factor was measured in a UV cross-linking experiment. Results Trans-acting factor interacting with the core sequence of GPEI (cGPEI) were found in human cervical adenocarcinoma cell line (HeLa) and rat hepatoma cell line (CBRH7919). These proteins were not expressed in normal rat liver. Although specific binding proteins that bound to GPEⅡ-1 were detected in all three cell types, a 64 kDa binding protein that exists in HeLa and CBRH7919 cells was absent in normal rat liver. Conclusion cGPEI, GPEII specific binding proteins expressed in HeLa and CBRH7919 cells may play an important role in the high transcriptional level of the rGSTP1 gene in tumor cells.
文摘Objective: To investigate the possible relationship between deletion of MTS/p16 gene and progression of endometrial carcinoma Methods: Forty six primary endometrial carcinoma, 7 tumor adjacent endometrial tissue, 10 normal endometrial tissue specimen and 5 xenografts from patients with endometrial carcinoma were examined for homozygous deletion of MTS/p16 gene by polymerase chain reaction based analysis Results: Of 46 endometrial cancer specimens, 9 showed homozygous deletion, no deletion was detected in the tumor adjacent and normal endometial tissues Nor was it detected in well differentiated endometrial carcinoma and all xenografts Conclusions: Deletion of MTS1/p16 gene might contribute to the progression of endometrial carcinoma and could be served as indicator for predicting prognosis
基金This work is supported by grant(HL32034) from the U.S.National Institute of Heart,Lung and Blood.
文摘Polycyclic aromatic hydrocarbons(PAHs),dimethyl beazanthracene(DMBA)and benzo-(a)-pyrene(BaP),stimulate the expression of the P1-450 gene in human lung tumor cells(ChaGo)in culture.A concentration-and time-dependent increase in the level of P1-450 specific mRNA sequences hasbeen observed in ChaGo cells treated with sublethal concentrations of DMBA.Results presentedsuggest that the parent compound causes the induction of P1-450 gene expression.The methylationpattern of the internal“-C-” residues of the “-CCGG-” sequence has been studied in a stretch ofabout 8000 base pairs(bp)of DNA sequence in and around the P1-450 gene in the control andDMBA-treated ChaGo cells.A comparative Southern blot analysis of the Msp Ⅰ/Hpa Ⅱ digestedDNA of the control and DMBA-treated ChaGo cells followed by hybridization with ^(32)P-labelledregion-specific probes,revealed that i)the internal “-C-” residues of the “-CCGG-” sequences ofdifferent regions of P1-450 gene were methylated to different degrees;ii)DMBA treatment of thecells induced hypomethylation of only the 5' end “-CCGG-” sequences of the gene;and iii)themethylation patterns of “-CCGG-” sequences of most of the coding region and of the 3' end regionof P1-450 gene were not affected by such DMBA treatment.
文摘Objective: To study the relationship between the expression of p27Kip1 and p53, and the infiltration, metastasis and prognosis in gastric carcinoma. Methods: The expression of p27Kip1 and p53 at protein level was determined by immunohistochemical assay (two-step method) in 100 cases of gastric carcinoma. Results: Of the 100 cases, the positive rate of p27Kip1 and p53 expressions were 44% and 49%, respectively. In the group of gastric carcinomas with deep infiltration (infiltration group), lymph nodes metastasis group (metastasis group) and death-within-5-years group (death group), the expression of p27Kip1 was statistically lower (_P<0.05). In the metastasis group and death group, the expression of p53 was significantly higher (P<0.05). The results of the monovariate analysis revealed that the 5-year survival rate of the high p27Kip1 expression group was 70.59%, which is higher than those of the low p27Kip1 expression group (54.55%) and the negative experession group (26%). The 5-year survival rate of the high p53 expression group was 19.23%, which was lower than those of the p53 low expression group (43.75%) and the negative group (53.19%). Cox multivariate analysis showed that p27Kip1, like p53, was an independent prognostic index. But p27Klpl protein expression was a stronger independent survival predictor (RR=3.06) than p53 expression (RR=2.33). Conclusion: The low expression of p27Kip1 and the high expression of p53 reflected the more frequent invasion and metastasis, which resulted in the reduced survival of patients. As an independent markers of the gastric carcinoma, the expression of p27Kip1 is more useful than that of p53 in the prognosis prediction of gastric carcinoma.
