Aug 05, 2024
Isolation and Transfection of Nicotiana benthamiana Mesophyll Protoplasts for Fluorescent Protein Visualization
- Namitha Nayak1,
- Sandhya Mehrotra1,
- Rajesh Mehrotra1
- 1BITS Pilani
Protocol Citation: Namitha Nayak, Sandhya Mehrotra, Rajesh Mehrotra 2024. Isolation and Transfection of Nicotiana benthamiana Mesophyll Protoplasts for Fluorescent Protein Visualization. protocols.io https://dx.doi.org/10.17504/protocols.io.e6nvw1my9lmk/v1
Manuscript citation:
Nayak N, Mehrotra R, Mehrotra S (2024) The N-region sequence
context impacts the chloroplast import efficiency of multi-TMD protein. Plant
molecular biology, 114(4), 88. https://doi.org/10.1007/s11103-024-01485-2
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: August 05, 2024
Last Modified: August 05, 2024
Protocol Integer ID: 104714
Keywords: Confocal microscopy, Fluorescent protein, Nicotiana benthamiana, Protoplast, Transfection, transfection of nicotiana benthamiana mesophyll protoplast, fluorescent protein visualization protoplast transfection, nicotiana benthamiana mesophyll protoplast, protoplasts from the leaf, fluorescent protein, better visualization of fluorescent protein, isolating protoplast, larger protoplast size, fluorescent, ephemeral arabidopsi, tagged protein, transfection efficiency, hardy plant
Funders Acknowledgements:
SERB
Grant ID: EMR/2016/002470
Abstract
Protoplast transfection is routinely used to study the sub-cellular localization of fluorescent-tagged proteins. Using N. benthamiana protoplasts is advantageous as it is a hardy plant that grows well, especially in tropical climates. Isolating protoplasts from the leaves of young 3-4-week-old N. benthamiana plants compensates for the benefits of using the ephemeral Arabidopsis. Moreover, the larger protoplast size of N. benthamiana offers better visualization of fluorescent proteins at lower magnifications. The protocol described here is an easy method for N. benthamiana protoplast isolation and transfection with simple and economical modifications to increase yield and transfection efficiency. The protocol is optimized for easy performance with minimal laboratory equipment.
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