TY  - JOUR
TI  - Sample Preparation and Fractionation of Arabidopsis thaliana Sperm and Vegetative Cell Nuclei by FACS
AU  - Chumak, Nina 
AU  - Mosiolek, Magdalena 
AU  - Schoft, Vera Karolina
VL  - 5
IS  - 22
PY  - 2015
DA  - 2015/11/20
SP  - e1664
C1  - Bio-protocol 2015;5:e1664
DO  - 10.21769/BioProtoc.1664
UR  - https://doi.org/10.21769/BioProtoc.1664
AB  - One of the major topics in plant and animal biology is sexual reproduction. It is, therefore, of great interest to isolate and study germ cells and accessory cells. The male gametophyte of the flowering plant Arabidopsis thaliana (A. thaliana), pollen, is the product of two post-meiotic mitotic divisions. Each mature pollen grain consists of two sperm cells contained within the vegetative cell, the non-reproductive companion cell. The tough pollen wall and its special nested structure make it difficult to study pollen cells separately. Here, we describe a simple and efficient method to fractionate A. thaliana sperm and vegetative cell nuclei by fluorescence activated cell sorting (FACS). Our protocol is based on differences in fluorescence intensity of sperm and vegetative cell nuclei stained with SYBR Green I. 100 plants yield about 1 x 106 sperm and 350,000 vegetative cell nuclei. This method can be used for purifying pollen nuclei of various A. thaliana wild-type accessions and mutant lines, and can, in principle, be adapted for pollen of other plant species.
KW  - SYBR Green
KW  - Fluorescence-activated cell sorting (FACS)
KW  - Arabidopsis
KW  - Pollen
KW  - Male gametophyte
JF  - Bio-protocol
SN  - 2331-8325
PB  - Bio-protocol LLC.
BIO101  - False