Cas9‐<scp>PF</scp>, an early flowering and visual selection marker system, enhances the frequency of editing event occurrence and expedites the isolation of genome‐edited and transgene‐free plants

Liu, Yong; Zeng, Jianmin; Cheng, Yuan; Guo, Yushuang; Yu, Haiqin; Li, Yongping; Huang, Changjun

doi:10.1111/pbi.13118

Cited by 39 publications

(21 citation statements)

References 10 publications

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“…In recent years, genome editing technologies, such as CRISPR/Cas9, have been used for functional genetic research in plant physiology and plant breeding (Ezura and Miura 2018;Yamamoto et al 2018b). For genome editing in plants, increasing the expression of genome-editing components may improve efficiency (Liu et al 2019). For example, transient expression of genome editing enzymes and guide RNA (gRNA) using the Tsukuba system improved base editing in tomatoes (Yuan et al 2021).…”

Section: Transient Expression System Applications Other Than Recombinant Protein Productionmentioning

confidence: 99%

Transient protein expression systems in plants and their applications

Nosaki

Hoshikawa

Ezura

et al. 2021

Plant Biotechnology

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The production of recombinant proteins is important in academic research to identify protein functions. Moreover, recombinant enzymes are used in the food and chemical industries, and high-quality proteins are required for diagnostic, therapeutic, and pharmaceutical applications. Though many recombinant proteins are produced by microbial or mammalian cell-based expression systems, plants have been promoted as alternative, cost-effective, scalable, safe, and sustainable expression systems. The development and improvement of transient expression systems have significantly reduced the period of protein production and increased the yield of recombinant proteins in plants. In this review, we consider the importance of plant-based expression systems for recombinant protein production and as genetic engineering tools.

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Section: Transient Expression System Applications Other Than Recombinant Protein Productionmentioning

confidence: 99%

Transient protein expression systems in plants and their applications

Nosaki

Hoshikawa

Ezura

et al. 2021

Plant Biotechnology

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“…Very recently, several examples have shown how the use of genome editing can also address the induction of early flowering, such as in kiwifruit (Varkonyi-Gasic et al, 2019). In tobacco the same approach was used to induce early flowering and precocious anthocyanin pigmentation (Liu et al, 2019). No published data are still available in citrus.…”

Section: Inducing Early Flowering As a Strategy To Shorten The Long Jmentioning

confidence: 99%

New Plant Breeding Techniques in Citrus for the Improvement of Important Agronomic Traits. A Review

et al. 2020

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New plant breeding techniques (NPBTs) aim to overcome traditional breeding limits for fruit tree species, in order to obtain new varieties with improved organoleptic traits and resistance to biotic and abiotic stress, and to maintain fruit quality achieved over centuries by (clonal) selection. Knowledge on the gene(s) controlling a specific trait is essential for the use of NPBTs, such as genome editing and cisgenesis. In the framework of the international scientific community working on fruit tree species, including citrus, NPBTs have mainly been applied to address pathogen threats. Citrus could take advantage of NPBTs because of its complex species biology (seedlessness, apomixis, high heterozygosity, and long juvenility phase) and aptitude for in vitro manipulation. To our knowledge, genome editing in citrus via transgenesis has successful for induced resistance to Citrus bacterial canker in sweet orange and grapefruit using the resistance gene CsLOB1. In the future, NPBTs will also be used to improve fruit traits, making them healthier. The regeneration of plants following the application of NPBTs is a bottleneck, making it necessary to optimize the efficiency of current protocols. The strengths and weaknesses of using explants from young in vitro plantlets, and from mature plants, will be discussed. Other major issues addressed in this review are related to the requirement for marker-free systems and shortening the long juvenility phase. This review aims to summarize methods and approaches available in the literature that are suitable to citrus, focusing on the principles observed before the use of NPBTs.

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“…For example, high-throughput fluorescence sorting of rice seeds has been developed to assist the screening of plants without transgenic fragments (Chang et al 2016). Moreover, some other natural color compounds can also be used as a marker to help select the transgene-free gene-edited plants (Liu et al 2019). Even though marker-assisted identification of transgene-free plants reduces workload by at least 75%, the strategy still requires significant amounts of labor and time.…”

Section: Fluorescence Marker-assisted Transgene Eliminationmentioning

confidence: 99%

“…Because the absence of any transgenes in gene-edited plants is a prerequisite for commercialization of any CRISPR-edited plants with stable valuable traits, several strategies have been employed to generate CRISPRedited plants that do not contain CRISPR constructs and other transgenes. The strategies include non-transgenic approaches (Luo et al 2015;Woo et al 2015), avoiding transgene integration (Chen et al 2018), maker-assisted tracking and selecting plants with/without transgenes (Gao et al 2016;He et al 2017;Liu et al 2019), and self-elimination of transgenes after plants are edited (He et al 2018(He et al , 2019. The goal is to generate transgene-free and gene-edited crop plants with greatly reduced costs of labor and time.…”

Section: Introductionmentioning

confidence: 99%

Technological breakthroughs in generating transgene-free and genetically stable CRISPR-edited plants

Zhao

2019

aBIOTECH

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CRISPR/Cas9 gene-editing technologies have been very effective in editing target genes in all major crop plants and offer unprecedented potentials in crop improvement. A major challenge in using CRISPR gene-editing technology for agricultural applications is that the target gene-edited crop plants need to be transgene free to maintain trait stability and to gain regulatory approval for commercial production. In this article, we present various strategies for generating transgene-free and target geneedited crop plants. The CRISPR transgenes can be removed by genetic segregation if the crop plants are reproduced sexually. Marker-assisted tracking and eliminating transgenes greatly decrease the time and labor needed for identifying the ideal transgene-free plants. Transgenes can be programed to undergo self-elimination when CRISPR genes and suicide genes are sequentially activated, greatly accelerating the isolation of transgene-free and target gene-edited plants. Transgene-free plants can also be generated using approaches that are considered non-transgenic such as ribonucleoprotein transfection, transient expression of transgenes without DNA integration, and nano-biotechnology. Here, we discuss the advantages and disadvantages of the various strategies in generating transgene-free plants and provide guidance for adopting the best strategies in editing a crop plant.

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Cas9‐PF, an early flowering and visual selection marker system, enhances the frequency of editing event occurrence and expedites the isolation of genome‐edited and transgene‐free plants

Cited by 39 publications

References 10 publications

Transient protein expression systems in plants and their applications

Transient protein expression systems in plants and their applications

New Plant Breeding Techniques in Citrus for the Improvement of Important Agronomic Traits. A Review

Technological breakthroughs in generating transgene-free and genetically stable CRISPR-edited plants

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