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Sensitive and high throughput quantification of abscisic acid based on quantitative real time immuno-PCR

文献类型： 外文期刊

作者： Su, Yi 2 ; Li, Wei 1 ; Huang, Zhigang 1 ; Wang, Ruozhong 1 ; Luo, Weigui 1 ; Liu, Qing 1 ; Tong, Jianhua 1 ; Xiao, Langta 1 ;

作者机构： 1.Hunan Agr Univ, Hunan Prov Key Lab Phytohormones & Growth Dev, Changsha, Hunan, Peoples R China

2.Hunan Agr Univ, Southern Reg Collaborat Innovat Ctr Grain & Oil C, Changsha, Hunan, Peoples R China

3.Hunan Acad Agr Sci, Tea Res Inst, Changsha 410125, Hunan, Peoples R China

关键词： qIPCR; ABA; Biotin; Avidin

期刊名称：PLANT METHODS （ 影响因子：4.993； 五年影响因子：5.312 ）

ISSN： 1746-4811

年卷期： 2018 年 14 卷

页码：

收录情况： SCI

摘要： Abscisic acid (ABA) functions as a stress phytohormone in many growth and developmental processes in plants. The ultra-sensitive determination of ABA would help to better understand its vital roles and action mechanisms. We report a new sensitive and high throughput quantitative real time immuno-PCR (qIPCR) method based on biotin-avidin linkage system for ABA determination in plants. ABA monoclonal antibody (McAb) coated on the inner surface of PCR well pretreated with glutaraldehyde. The pre-prepared probe complex, including biotinylated McAb, biotinylated DNA and streptavidin linker, was convenient for high throughput operations. Finally, probe DNA was quantified by real-time PCR. The detectable ranges were from 10 to 40 ng/L with a limit of detection (LOD) of 2.5 fg. ABA contents in plant sample were simultaneously analyzed using LC-MS/MS to validate the qIPCR method. The results showed that qIPCR method has good specificity and repeatability with a recovery rate of 96.9%. The qIPCR method is highly sensitive for ABA quantification for actual plant samples with an advantage of using crude extracts instead of intensively purified samples.

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