【摘要】：用RT-PCR法分別擴(kuò)增和分離出CTN株和SRV9株狂犬病毒糖蛋白部分表位基因,構(gòu)建真核表達(dá)質(zhì)粒pcDNA/CTN和pcDNA/SRV9CTN,提質(zhì)粒酶切鑒定并測(cè)序。堿裂解法大提質(zhì)粒, Sepharose 4FF柱層析純化,肌肉注射免疫NIH小鼠, ELISA法檢測(cè)該DNA疫苗誘導(dǎo)小鼠產(chǎn)生抗狂犬病毒的體液免疫效果。小鼠脾淋巴細(xì)胞轉(zhuǎn)化實(shí)驗(yàn)和CTLL-2細(xì)胞株/MTT法測(cè)定IL-2活性評(píng)估其誘導(dǎo)的細(xì)胞免疫效果。 ELISA法檢測(cè)到抗狂犬病毒抗體pcDNA/CTN和pcDNA/SRV9CTN的OD450分別為0.426和0.384,在誘導(dǎo)的細(xì)胞免疫效果中,質(zhì)粒pcDNA/CTN DNA疫苗免疫組ConA刺激指數(shù)均值為7.140±0.5613,而pcDNA/SRV9CTN和空載體對(duì)照組則分別為6.476±0.5436和1.786±0.528。在免疫小鼠血清中的IL-2活性測(cè)定方面,質(zhì)粒pcDNA/CTN免疫組達(dá)到1.812±0.068 IU/ml,而pcDNA/SRV9CTN和空載體對(duì)照免疫組則分別為1.614±0.062IU/ml和0.163±0.031IU/ml。pcDNA/CTN DNA疫苗、pcDNA/SRV9CTN DNA疫苗和空載體免疫小鼠的保護(hù)率分別為75%、65%和25%,其差異具有顯著性。
[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 鹵0.5613. PcDNA/SRV9CTN and empty carrier control group were 6.476 鹵0.5436 and 1.786 鹵0.528 respectively. The activity of IL-2 in serum of immunized mice was 1.812 鹵0.068 IU/ml, in plasmid pcDNA/CTN immunized group. The protective rates of pcDNA/SRV9CTN and empty vector immunized mice were 1.614 鹵0.062IU/ml and 0.163 鹵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.
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2006
【分類號(hào)】：R392

【參考文獻(xiàn)】

相關(guān)期刊論文 前10條

1 王三虎,袁慧君;狂犬病病毒新型疫苗研究進(jìn)展[J];河南畜牧獸醫(yī);2004年11期

2 吳小紅,劉景華,董關(guān)木;中國(guó)狂犬病疫苗CTN-1-V株糖蛋白基因的克隆及序列分析[J];中國(guó)生物制品學(xué)雜志;2003年06期

3 張玉慧,高春潤(rùn),王忠海,趙鋼,孫英,李春孝,王世龍,秦賢,俞永新;地鼠腎細(xì)胞培養(yǎng)的CTN株狂犬病新疫苗研究[J];微生物學(xué)免疫學(xué)進(jìn)展;2002年04期

4 周志軍,朱華松,李健,秦麗,李軍,孫文,黃貴彪;不同狂犬病毒株抗原反應(yīng)性的比較研究[J];微生物學(xué)免疫學(xué)進(jìn)展;2005年01期

5 楊裕華,汪天虹,解力,趙世立;狂犬病毒糖蛋白主要免疫活性位點(diǎn)的質(zhì)粒載體構(gòu)建[J];現(xiàn)代預(yù)防醫(yī)學(xué);2000年04期

6 李萍,嚴(yán)家新,朱家鴻;狂犬病毒糖蛋白DNA疫苗的研制及其免疫效果的觀察[J];中國(guó)病毒學(xué);1997年02期

7 唐青,Lillian A Orciari,Charles E.Rupprechti,趙秀芹,李志剛,楊為松;我國(guó)四株狂犬病毒糖蛋白基因序列分析和位點(diǎn)比較[J];中國(guó)病毒學(xué);2000年01期

8 張永振;中國(guó)狂犬病的流行病學(xué)[J];中國(guó)計(jì)劃免疫;2005年02期

9 唐青,李浩;中國(guó)狂犬病流行近況及相關(guān)因素分析[J];中華流行病學(xué)雜志;2005年03期

10 史秀山;轉(zhuǎn)移因子增強(qiáng)狂犬病疫苗免疫效果的研究[J];中國(guó)人獸共患病雜志;2004年11期

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