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A high-throughput transient expression system for rice

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A high-throughput transient expression system for rice. / Page, Michael; Parry, Martin Afan John; Carmo-Silva, Ana Elizabete.

In: Plant, Cell and Environment, Vol. 42, No. 7, 01.07.2019, p. 2057-2064.

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@article{e37ef2c663cc4257bb3483adeaafca7d,
title = "A high-throughput transient expression system for rice",
abstract = "Rice is an important global crop and represents a vital source of calories for many food insecure regions. Efforts to improve this crop by improving yield, nutritional content, stress tolerance, or resilience to climate change are certain to include biotechnological approaches, which rely on the expression of transgenes in planta. The throughput and cost of currently available transgenic expression systems is frequently incompatible with modern, high-throughput molecular cloning methods. Here, we present a protocol for isolating high yields of green rice protoplasts and for PEG-mediated transformation of isolated protoplasts. Factors affecting transformation efficiency were investigated, and the resulting protocol is fast, cheap, robust, high-throughput, and does not require specialist equipment. When coupled to a high-throughput modular cloning system such as Golden Gate, this transient expression system provides a valuable resource to help break the {"}design-build-test{"} bottleneck by permitting the rapid screening of large numbers of transgenic expression cassettes prior to stable plant transformation. We used this system to rapidly assess the expression level, subcellular localisation, and protein aggregation pattern of nine single-gene expression cassettes, which represent the essential component parts of the beta-cyanobacterial carboxysome.",
keywords = "Rice, Oryza sativa, protoplasts, transient expression, confocal microscopy, cellular localisation, high-throughput, synthetic biology, Golden Gate, carboxysome",
author = "Michael Page and Parry, {Martin Afan John} and Carmo-Silva, {Ana Elizabete}",
year = "2019",
month = jul
day = "1",
doi = "10.1111/pce.13542",
language = "English",
volume = "42",
pages = "2057--2064",
journal = "Plant, Cell and Environment",
issn = "0140-7791",
publisher = "Wiley",
number = "7",

}

RIS

TY - JOUR

T1 - A high-throughput transient expression system for rice

AU - Page, Michael

AU - Parry, Martin Afan John

AU - Carmo-Silva, Ana Elizabete

PY - 2019/7/1

Y1 - 2019/7/1

N2 - Rice is an important global crop and represents a vital source of calories for many food insecure regions. Efforts to improve this crop by improving yield, nutritional content, stress tolerance, or resilience to climate change are certain to include biotechnological approaches, which rely on the expression of transgenes in planta. The throughput and cost of currently available transgenic expression systems is frequently incompatible with modern, high-throughput molecular cloning methods. Here, we present a protocol for isolating high yields of green rice protoplasts and for PEG-mediated transformation of isolated protoplasts. Factors affecting transformation efficiency were investigated, and the resulting protocol is fast, cheap, robust, high-throughput, and does not require specialist equipment. When coupled to a high-throughput modular cloning system such as Golden Gate, this transient expression system provides a valuable resource to help break the "design-build-test" bottleneck by permitting the rapid screening of large numbers of transgenic expression cassettes prior to stable plant transformation. We used this system to rapidly assess the expression level, subcellular localisation, and protein aggregation pattern of nine single-gene expression cassettes, which represent the essential component parts of the beta-cyanobacterial carboxysome.

AB - Rice is an important global crop and represents a vital source of calories for many food insecure regions. Efforts to improve this crop by improving yield, nutritional content, stress tolerance, or resilience to climate change are certain to include biotechnological approaches, which rely on the expression of transgenes in planta. The throughput and cost of currently available transgenic expression systems is frequently incompatible with modern, high-throughput molecular cloning methods. Here, we present a protocol for isolating high yields of green rice protoplasts and for PEG-mediated transformation of isolated protoplasts. Factors affecting transformation efficiency were investigated, and the resulting protocol is fast, cheap, robust, high-throughput, and does not require specialist equipment. When coupled to a high-throughput modular cloning system such as Golden Gate, this transient expression system provides a valuable resource to help break the "design-build-test" bottleneck by permitting the rapid screening of large numbers of transgenic expression cassettes prior to stable plant transformation. We used this system to rapidly assess the expression level, subcellular localisation, and protein aggregation pattern of nine single-gene expression cassettes, which represent the essential component parts of the beta-cyanobacterial carboxysome.

KW - Rice

KW - Oryza sativa

KW - protoplasts

KW - transient expression

KW - confocal microscopy

KW - cellular localisation

KW - high-throughput

KW - synthetic biology

KW - Golden Gate

KW - carboxysome

U2 - 10.1111/pce.13542

DO - 10.1111/pce.13542

M3 - Journal article

VL - 42

SP - 2057

EP - 2064

JO - Plant, Cell and Environment

JF - Plant, Cell and Environment

SN - 0140-7791

IS - 7

ER -