Development of a Rapid and Efficient Protoplast Isolation and Transfection Method for Chickpea (Cicer arietinum)

Authors

Ninghui Cheng
Paul A Nakata

Language

English

Publication Date

1-1-2020

Journal

MethodsX

DOI

10.1016/j.mex.2020.101025

PMID

32874941

PMCID

PMC7452273

PubMedCentral® Posted Date

8-8-2020

PubMedCentral® Full Text Version

Post-print

Abstract

Chickpea (Cicer arietinum L.) is the second most important grain legume worldwide. Recent advances in the sequencing of the chickpea genome has provided a new and valuable resource to aid efforts in gene discovery and crop trait improvement. Technical difficulties in stable chickpea transgenics and the lack of a transient expression system for rapid analysis of gene expression and function; however, has limited the usefulness of this genomic resource. As a step toward alleviating this limitation, we report here the development of a simple and efficient transient gene expression protocol. Using leaves from chickpea seedlings, we have established a procedure that enables the generation of large quantities of vital chickpea protoplasts within only a few hours. In addition, we have optimized a PEG-calcium-mediated transfection method to efficiently deliver exogenous DNA into the chickpea protoplast. The current study is the first to present a detailed step-by-step procedures for protoplast isolation, evaluation, transfection, and application in chickpea. In addition, we optimize the transfection efficiency which has not been previously reported. Our protoplast transfection approach provides a platform that will allow rapid high-throughput screening and systematic characterization of gene expression and function. Knowledge gained through such studies will benefit current efforts to improve chickpea production and quality.

• Modified enzymatic digestion solution for higher yield and viability.
• Optimize transfection of chickpea protoplasts.

Keywords

Protoplast, Transfection, Chickpea, Transient expression, Subcellular localization

Published Open-Access

yes

Recommended Citation

Ninghui Cheng and Paul A Nakata, "Development of a Rapid and Efficient Protoplast Isolation and Transfection Method for Chickpea (Cicer arietinum)" (2020). Children’s Nutrition Research Center Staff Publications. 85.
https://digitalcommons.library.tmc.edu/staff_pub/85