【摘要】：用RT-PCR法分別擴增和分離出CTN株和SRV9株狂犬病毒糖蛋白部分表位基因,構建真核表達質粒pcDNA/CTN和pcDNA/SRV9CTN,提質粒酶切鑒定并測序。堿裂解法大提質粒, Sepharose 4FF柱層析純化,肌肉注射免疫NIH小鼠, ELISA法檢測該DNA疫苗誘導小鼠產生抗狂犬病毒的體液免疫效果。小鼠脾淋巴細胞轉化實驗和CTLL-2細胞株/MTT法測定IL-2活性評估其誘導的細胞免疫效果。 ELISA法檢測到抗狂犬病毒抗體pcDNA/CTN和pcDNA/SRV9CTN的OD450分別為0.426和0.384,在誘導的細胞免疫效果中,質粒pcDNA/CTN DNA疫苗免疫組ConA刺激指數均值為7.140±0.5613,而pcDNA/SRV9CTN和空載體對照組則分別為6.476±0.5436和1.786±0.528。在免疫小鼠血清中的IL-2活性測定方面,質粒pcDNA/CTN免疫組達到1.812±0.068 IU/ml,而pcDNA/SRV9CTN和空載體對照免疫組則分別為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.
【學位授予單位】：吉林大學
【學位級別】：碩士
【學位授予年份】：2006
【分類號】：R392

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