狂犬病毒糖蛋白DNA疫苗的构建及其诱导小鼠的免疫应答
发布时间：2019-01-16 04:20
【摘要】：用RT-PCR法分别扩增和分离出CTN株和SRV9株狂犬病毒糖蛋白部分表位基因,构建真核表达质粒pcDNA/CTN和pcDNA/SRV9CTN,提质粒酶切鉴定并测序。碱裂解法大提质粒, Sepharose 4FF柱层析纯化,肌肉注射免疫NIH小鼠, ELISA法检测该DNA疫苗诱导小鼠产生抗狂犬病毒的体液免疫效果。小鼠脾淋巴细胞转化实验和CTLL-2细胞株/MTT法测定IL-2活性评估其诱导的细胞免疫效果。 ELISA法检测到抗狂犬病毒抗体pcDNA/CTN和pcDNA/SRV9CTN的OD450分别为0.426和0.384,在诱导的细胞免疫效果中,质粒pcDNA/CTN DNA疫苗免疫组ConA刺激指数均值为7.140±0.5613,而pcDNA/SRV9CTN和空载体对照组则分别为6.476±0.5436和1.786±0.528。在免疫小鼠血清中的IL-2活性测定方面,质粒pcDNA/CTN免疫组达到1.812±0.068 IU/ml,而pcDNA/SRV9CTN和空载体对照免疫组则分别为1.614±0.062IU/ml和0.163±0.031IU/ml。pcDNA/CTN DNA疫苗、pcDNA/SRV9CTN DNA疫苗和空载体免疫小鼠的保护率分别为75%、65%和25%,其差异具有显著性。
[Abstract]:Some epitopes of rabies virus glycoprotein of CTN strain and SRV9 strain were amplified and isolated by RT-PCR method. The eukaryotic expression plasmids pcDNA/CTN and pcDNA/SRV9CTN, were digested and sequenced. The plasmid was extracted by alkaline lysis and purified by Sepharose 4FF column chromatography. NIH mice were immunized by intramuscular injection. The humoral immunity of mice induced by the DNA vaccine against rabies virus was detected by ELISA method. Spleen lymphocyte transformation test and CTLL-2 cell line / MTT assay were used to evaluate the effect of IL-2. The OD450 of anti-rabies virus antibody pcDNA/CTN and pcDNA/SRV9CTN detected by ELISA method were 0.426 and 0.384, respectively. The average ConA stimulation index of plasmid pcDNA/CTN DNA vaccine immunization group was 7.140 卤0.5613. PcDNA/SRV9CTN and empty carrier control group were 6.476 卤0.5436 and 1.786 卤0.528 respectively. The activity of IL-2 in serum of immunized mice was 1.812 卤0.068 IU/ml, in plasmid pcDNA/CTN immunized group. The protective rates of pcDNA/SRV9CTN and empty vector immunized mice were 1.614 卤0.062IU/ml and 0.163 卤0.031IU/ml.pcDNA/CTN DNA, respectively. The protective rates of pcDNA/SRV9CTN DNA vaccine and empty vector immunized mice were 75% and 25%, respectively. The difference was significant.
【学位授予单位】：吉林大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R392
本文编号：2409455
[Abstract]:Some epitopes of rabies virus glycoprotein of CTN strain and SRV9 strain were amplified and isolated by RT-PCR method. The eukaryotic expression plasmids pcDNA/CTN and pcDNA/SRV9CTN, were digested and sequenced. The plasmid was extracted by alkaline lysis and purified by Sepharose 4FF column chromatography. NIH mice were immunized by intramuscular injection. The humoral immunity of mice induced by the DNA vaccine against rabies virus was detected by ELISA method. Spleen lymphocyte transformation test and CTLL-2 cell line / MTT assay were used to evaluate the effect of IL-2. The OD450 of anti-rabies virus antibody pcDNA/CTN and pcDNA/SRV9CTN detected by ELISA method were 0.426 and 0.384, respectively. The average ConA stimulation index of plasmid pcDNA/CTN DNA vaccine immunization group was 7.140 卤0.5613. PcDNA/SRV9CTN and empty carrier control group were 6.476 卤0.5436 and 1.786 卤0.528 respectively. The activity of IL-2 in serum of immunized mice was 1.812 卤0.068 IU/ml, in plasmid pcDNA/CTN immunized group. The protective rates of pcDNA/SRV9CTN and empty vector immunized mice were 1.614 卤0.062IU/ml and 0.163 卤0.031IU/ml.pcDNA/CTN DNA, respectively. The protective rates of pcDNA/SRV9CTN DNA vaccine and empty vector immunized mice were 75% and 25%, respectively. The difference was significant.
【学位授予单位】：吉林大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R392
【参考文献】
相关期刊论文 前10条
1 王三虎,袁慧君;狂犬病病毒新型疫苗研究进展[J];河南畜牧兽医;2004年11期
2 吴小红,刘景华,董关木;中国狂犬病疫苗CTN-1-V株糖蛋白基因的克隆及序列分析[J];中国生物制品学杂志;2003年06期
3 张玉慧,高春润,王忠海,赵钢,孙英,李春孝,王世龙,秦贤,俞永新;地鼠肾细胞培养的CTN株狂犬病新疫苗研究[J];微生物学免疫学进展;2002年04期
4 周志军,朱华松,李健,秦丽,李军,孙文,黄贵彪;不同狂犬病毒株抗原反应性的比较研究[J];微生物学免疫学进展;2005年01期
5 杨裕华,汪天虹,解力,赵世立;狂犬病毒糖蛋白主要免疫活性位点的质粒载体构建[J];现代预防医学;2000年04期
6 李萍,严家新,朱家鸿;狂犬病毒糖蛋白DNA疫苗的研制及其免疫效果的观察[J];中国病毒学;1997年02期
7 唐青,Lillian A Orciari,Charles E.Rupprechti,赵秀芹,李志刚,杨为松;我国四株狂犬病毒糖蛋白基因序列分析和位点比较[J];中国病毒学;2000年01期
8 张永振;中国狂犬病的流行病学[J];中国计划免疫;2005年02期
9 唐青,李浩;中国狂犬病流行近况及相关因素分析[J];中华流行病学杂志;2005年03期
10 史秀山;转移因子增强狂犬病疫苗免疫效果的研究[J];中国人兽共患病杂志;2004年11期
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