Nuclear DNA and protein content evaluation in Taxus plant cell cultures using multiparameter flow cytometry

Abstract

Plant cell cultures of Taxus provide the most reliable production methods for the anti-cancer drug paclitaxel. In order to comprehend the inherent culture heterogeneity and production variability in cell cultures, it is essential that the cellular metabolism is studied at the genomic level. Genomic stability in plant cell cultures is crucial as it affects cell growth and division, metabolite accumulation and protein synthesis. A rapid and efficient method to prepare nuclei suspensions from aggregated cell cultures of Taxus was employed. Methods were subsequently developed to simultaneously stain them for DNA and protein content using Propidium Iodide and Fluorescein Isothiocyanate respectively. Flow cytometry was used to analyze and quantify the DNA content and genome size of Taxus using known reference species as standards. Furthermore, their genomic stability was evaluated by correlating DNA content and genome size with cell size and complexity, protein content, and elicitation effects using multiparameter flow cytometry. These techniques to evaluate and correlate various culture characteristics can be very useful in designing superior bio processes for enhanced production.


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