Improving agroinfiltration-based transient gene expression in Nicotiana benthamiana

Plant Methods
Karlah NorkunasBenjamin Dugdale

Abstract

Agroinfiltration is a simple and effective method of delivering transgenes into plant cells for the rapid production of recombinant proteins and has become the preferred transient expression platform to manufacture biologics in plants. Despite its popularity, few studies have sought to improve the efficiency of agroinfiltration to further increase protein yields. This study aimed to increase agroinfiltration-based transient gene expression in Nicotiana benthamiana by improving all levels of transgenesis. Using the benchmark pEAQ-HT deconstructed virus vector system and the GUS reporter enzyme, physical, chemical, and molecular features were independently assessed for their ability to enhance Agrobacterium-mediated transformation and improve protein production capacities. Optimal Agrobacterium strain, cell culture density and co-cultivation time for maximal transient GUS (β-glucuronidase) expression were established. The effects of chemical additives in the liquid infiltration media were investigated and acetosyringone (500 μM), the antioxidant lipoic acid (5 μM), and a surfactant Pluronic F-68 (0.002%) were all shown to significantly increase transgene expression. Gene products known to suppress post-transcriptional gene silenc...Continue Reading

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Citations

Mar 29, 2020·Frontiers in Plant Science·Ramona PersadScott C Lenaghan
Jul 10, 2020·Molecular Biology Reports·Behnam SedaghatiMojgan Bandehpour
Aug 11, 2019·Plant Molecular Biology·Tong ZhangSixue Chen
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Nov 17, 2020·Frontiers in Plant Science·Daniel TuséJohannes Felix Buyel
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Aug 10, 2021·The Plant Journal : for Cell and Molecular Biology·Philippe V JutrasRenier A L van der Hoorn

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Methods Mentioned

BETA
transgenic
PCR
RepA
ELISA

Software Mentioned

HT
Excel
pEAQ
pNEEDS

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