Chapter 6: An Improved Leaf Protoplast System for Highly Efficient Transient Expression in Switchgrass (Panicum virgatum L.)

Chien-Yuan Lin, Hui Wei, Bryon Donohoe, Melvin Tucker, Michael Himmel

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

1 Scopus Citations

Abstract

As a robust perennial C4-type monocot plant and a native species to North America, switchgrass (Panicum virgatum) has been evaluated and designated as a strong candidate bioenergy crop by the U.S. DOE. Although genetic modifications of switchgrass have been used to successfully reduce the recalcitrance of switchgrass biomass for biofuel production, the generation of transgenic switchgrass is still a slow and laborious process. A transient protoplast system can provide an excellent platform to accelerate the selection of genes-of-interest for tailoring switchgrass biomass. However, partially due to the lack of the complete genomic information, the attempts to optimize the transient protoplast system for switchgrass remain scarce. In this chapter, we provide an improved protocol for switchgrass protoplast isolation, increased transformation efficiency using CsCl gradient ultracentrifugation-derived plasmid DNA and extended application of the transient switchgrass protoplast system to analyze protein expression using western blot.

Original languageAmerican English
Title of host publicationMetabolic Pathway Engineering
Subtitle of host publicationMethods in Molecular Biology, Volume 2096
EditorsM. E. Himmel, Y. J. Bomble
PublisherHumana Press Inc.
Pages61-79
Number of pages19
DOIs
StatePublished - 2020

Publication series

NameMethods in Molecular Biology
Volume2096
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

Bibliographical note

Publisher Copyright:
© Springer Science+Business Media, LLC, part of Springer Nature 2020.

NREL Publication Number

  • NREL/CH-2700-74951

Keywords

  • CsCl gradient ultracentrifugation
  • GFP expression
  • Protoplast
  • Switchgrass
  • Transient expression system

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