Open AccessDevelopment of gene expression system in egg cells and zygotes isolated from rice and maize
Author(s) -
Koiso Narumi,
Toda Erika,
Ichikawa Masako,
Kato Norio,
Okamoto Takashi
Publication year - 2017
Publication title -
plant direct
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.211
H-Index - 11
ISSN - 2475-4455
DOI - 10.1002/pld3.10
Subject(s) - transfection , zygote , microbiology and biotechnology , biology , green fluorescent protein , plasmid , exogenous dna , peg ratio , reporter gene , gene , gene delivery , electroporation , gene expression , embryo , genetics , embryogenesis , finance , economics
Polyethylene glycol calcium ( PEG ‐Ca 2+ ) transfection‐mediated analysis allows rapid and efficient examination of gene function. To investigate the diverse cellular functions of genes of interest in plant cells, macromolecules, such as DNA , RNA , and proteins, are delivered into protoplasts prepared from somatic tissues or calli using a PEG ‐Ca 2+ transfection procedure. To take advantage of this macromolecule delivery system in the reproductive and developmental biology of angiosperms, this study established a PEG ‐Ca 2+ transfection system with isolated egg cells and zygotes. The conditions for PEG and plasmid DNA concentrations for transfection of rice egg cells were first addressed, and ~30% of PEG ‐Ca 2+ ‐transfected egg cells showed exogenous and transient expressions of fluorescent proteins from plasmid DNA delivered into the cells. Interestingly, a dual expression of two different fluorescent proteins in the same egg cell using two kinds of plasmid DNA s was also observed. For PEG ‐Ca 2+ transfection with maize zygotes, ~80% of zygotes showed expression of GFP proteins from plasmid DNA . Importantly, PEG ‐transfected zygotes developed normally into cell masses and mature plants. These results suggest that the present PEG ‐Ca 2+ ‐mediated transient expression system provides a novel and effective platform for expressing and analyzing genes of interest in egg cells and zygotes. Moreover, combined with the CRISPR /Cas9 approach, the present transient expression system in zygotes will become a powerful and alternative tool for the preparation of gene‐edited plants.