以口蹄疫病毒株AFT2 RNA为模板,反转录并扩增3D聚合酶基因,PCR纯化产物与pGEM-T easy载体连接并转化JM109菌株,对经凝胶电泳、PCR和EcoRI酶切法鉴定为阳性的重组质粒进行测序,通过序列比对获得AF72 3D聚合酶的核苷酸序列和推导氨基酸序列,综合分析3D聚合酶的亲水性、可塑性、抗原指数以及表面可能性等参数,预测其潜在B细胞抗原表位并人工合成表位肽段,利用间接ELISA对潜在表位进行筛选鉴定,结果显示,表位3D2和3D4为病毒株AFT2 3D聚合酶的优势B细胞表位,该结果将为进一步的FMDV多表位疫苗研究奠定基础.
张昱
,
王永录
,
张永光
,
潘丽
,
方玉珍
,
刘力宽
,
蒋守田
,
吕建亮
,
张中旺
,
张淑刚
,
李正丰
,
杜进鑫
. 口蹄疫病毒3D聚合酶B细胞表位的筛选鉴定[J]. 华北农学报, 2009
, 24(3)
: 69
-73
.
DOI: 10.7668/hbnxb.2009.03.015
Foot2and2mouth disease virus strain AF72 RNAs were used as templates for RT2PCR to amplify the 3D gene. The purified PCR products were cloned into pGEm2T easy vectors and transformed into E. coli JM109. The positive recombinant plasmids identified by electrophoresis,PCR,and EcoR Ⅰ cleavage were sequenced. The nucleotide and amino acid sequence were obtained by comparing with the full2length sequence of the other reference strains. Potential B cell epi2 topes of 3D were predicted and epitope peptide segments were synthesized. They were identified by indirect ELISA. The re2 sult showed that 3D2 and 3D4 were predominant B cell epitope of 3D,it provided a basisfor further studyon the multi2epi2 tope vaccine of FMD.
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