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

BACKGROUND: 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. RESULTS: We report a new sensitive and high throughput quantitative...

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Detalles Bibliográficos
Autores principales: Su, Yi, Li, Wei, Huang, Zhigang, Wang, Ruozhong, Luo, Weigui, Liu, Qing, Tong, Jianhua, Xiao, Langtao
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6260876/
https://www.ncbi.nlm.nih.gov/pubmed/30534191
http://dx.doi.org/10.1186/s13007-018-0371-y
Descripción
Sumario:BACKGROUND: 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. RESULTS: 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%. CONCLUSION: 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.