期刊
GENETICS AND MOLECULAR RESEARCH
卷 12, 期 4, 页码 4371-4382出版社
FUNPEC-EDITORA
DOI: 10.4238/2013.October.10.3
关键词
Alkaline phosphatase; pET-32a expression vector; ELISA; Helicoverpa armigera Hub.; Polyclonal antibody
资金
- Ministry of Science and Technology of China [200903052]
- Ministry of Agriculture, China
- '13115' Sci-Tech Innovation Project of Shaanxi Province [2007ZDKG-14]
Alkaline phosphatase (ALP) of Helicoverpa armigera Hub. (Lepidoptera; Noctuidae) (GenBank accession No. EU729322) was cloned and expressed. The target gene H.alpha-ALP, having an open reading frame of 1608 bp, was reverse-transcribed from cDNA by the polymerase chain reaction. The open reading frame of the target gene was cloned into the pET-32a expression vector to obtain recombinant protein in Escherichia coli DE-3 cells for the subsequent production of polyclonal antibody. New Zealand white rabbits were used for production of anti-pET-32a-H. a-ALP. The production of antibody was also optimized by employing ELISA for titer determination. The produced antiserum was processed and used as an antibody. Western blot results showed that the polyclonal antibody produced was capable of effectively binding target protein not only from H. armigera but also from other lepidopterans such as Mythimna separata and Plutella xylostella. This antibody was also used to detect levels of ALP within different instars of H. armigera. Thus, it is concluded that this antibody-based assay is very useful for the effective detection of gene-specific expression. Furthermore, it may also be used to detect the expression levels and tissue localization of ALP, as well as in other physiological studies involving this enzyme.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据