Improved and Reproducible Flow Cytometry Methodology for Nuclei Isolation from Single Root Meristem

Root meristems have increasingly been target of cell cycle studies by flow cytometric DNA content quantification. Moreover, roots can be an alternative source of nuclear suspension when leaves become unfeasible and for chromosome analysis and sorting. In the present paper, a protocol for intact nucl...

Full description

Saved in:
Bibliographic Details
Published inJournal of botany Vol. 2010; no. 2010; pp. 1 - 7
Main Authors Silva, Thaís Cristina Ribeiro, Abreu, Isabella Santiago, Carvalho, Carlos Roberto
Format Journal Article
LanguageEnglish
Published Cairo, Egypt Hindawi Puplishing Corporation 01.01.2010
Hindawi Publishing Corporation
Hindawi Limited
Subjects
Chromosomes
Flow cytometry
Studies
Online AccessGet full text

Cover

More Information
Summary:Root meristems have increasingly been target of cell cycle studies by flow cytometric DNA content quantification. Moreover, roots can be an alternative source of nuclear suspension when leaves become unfeasible and for chromosome analysis and sorting. In the present paper, a protocol for intact nuclei isolation from a single root meristem was developed. This proceeding was based on excision of the meristematic region using a prototypical slide, followed by short enzymatic digestion and mechanical isolation of nuclei during homogenization with a hand mixer. Such parameters were optimized for reaching better results. Satisfactory nuclei amounts were extracted and analyzed by flow cytometry, producing histograms with reduced background noise and CVs between 3.2 and 4.1%. This improved and reproducible technique was shown to be rapid, inexpensive, and simple for nuclear extraction from a single root tip, and can be adapted for other plants and purposes.
ISSN:2090-0120
2090-0139
DOI:10.1155/2010/320609