Molecular cloning and characterization of Babesia orientalis rhoptry-associated protein 1
Molecular cloning and characterization of Babesia orientalis rhoptry-associated protein 1
复制标题
东方巴贝虫菱形相关蛋白 1 的分子克隆和表征
DOI：
10.1016/j.vetpar.2014.08.007
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发表时间：
2014-10-15
影响因子：
2.6
通讯作者：
Zhao, Jun-Long
中科院分区：
文献类型：
--
作者：
Yu, Qian;He, Lan;Zhao, Jun-Long
通过B免疫筛选，从东方Babelioralis的cDNA表达文库中获得了东方Babelioralis的棒状体相关蛋白1（RAP-1）基因。东方蜱感染的布法罗血清。该基因全长1732 bp，开放阅读框（ORF）为1434 bp，编码478个氨基酸，分子量为52.5 kDa。将ORF克隆到pGEX-KG质粒中，随后表达为GST融合蛋白。重组RAP-1 B.纯化东方红细胞（rBoRAP-1），并使用Western印迹法评估其作为抗原。天然BoRAP-1被兔中产生的针对rBoRAP-1的抗体识别。在感染B的红细胞中观察到强免疫荧光信号。东方系统发育分析表明，B.东方巴贝属属于巴贝属分支，与牛巴贝属和羊巴贝属亲缘关系最近，这与以往报道的基于18 SrRNA和HSP 70基因的分类结果相似。本研究提示BoRAP-1可能是一种潜在的诊断抗原，RAP-1基因可用于巴氏杆菌和泰勒虫的分类。(C)© 2014 Elsevier B. V.保留所有权利。
The rhoptry-associated protein 1 (RAP-1) gene of Babesia orientalis was obtained from a cDNA expression library by immunoscreening with B. orientalis-infected water buffalo sera. The nucleotide sequence of the cDNA was 1732 bp with an open reading frame (ORF) of 1434 bp, encoding a polypeptide of 478 amino acid residues with a predicted size of 52.5 kDa. The ORF was cloned into a pGEX-KG plasmid and subsequently expressed as a GST-fusion protein. The recombinant RAP-1 of B. orientalis (rBoRAP-1) was purified and evaluated as an antigen using Western blotting. The native BoRAP-1 was recognized by the antibodies raised in rabbits against rBoRAP-1. Strong immunofluorescence signals were observed in erythrocytes infected with B. orientalis. Phylogentic analysis revealed that B. orientalis fell into a Babesia clade and most closely related to Babesia bovis and Babesia ovis, which was similar to the previous reported trees based on 18S rRNA and HSP70 genes. The present study suggests that the BoRAP-1 might be a potential diagnostic antigen, and the RAP-1 genes can aid in the classification of Babesia and Theileria species. (C) 2014 Elsevier B.V. All rights reserved.