High-Efficiency Genome Editing in Arabidopsis Using YAO Promoter-Driven CRISPR/Cas9 System

Yan, Liuhua; Wei, Shaowei; Wu, Yaorong; Hu, Ruofei; Li, Hong-Ju; Yang, Wei‐Cai; Xie, Qi

doi:10.1016/j.molp.2015.10.004

Cited by 355 publications

(280 citation statements)

References 14 publications

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“…We have also included the Arabidopsis Ec1.2 and YAO promoters, which are egg cell and embryo sac/pollen specific, respectively. Although Arabidopsis was not our target species in this study, these two promoters have been shown to enhance mutagenesis in Arabidopsis when used to drive Cas9 expression Yan et al, 2015). In our toolkit, these promoters can drive either Cas9 expression or the polycistronic gRNA arrays.…”

Section: Discussionmentioning

confidence: 99%

“…For example, indel frequencies have been increased by coupling SSNs with exonucleases (Certo et al, 2012;Kwon et al, 2012), altering guide RNA (gRNA) architecture or using specialized promoters Yan et al, 2015). Increased specificity has been achieved using paired TALEN or CRISPR/Cas9 nickases , truncated gRNAs (Fu et al, 2014), or dimeric CRISPR/Cas9 nucleases (Guilinger et al, 2014;Tsai et al, 2014).…”

Section: Introductionmentioning

confidence: 99%

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A Multipurpose Toolkit to Enable Advanced Genome Engineering in Plants

et al. 2017

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We report a comprehensive toolkit that enables targeted, specific modification of monocot and dicot genomes using a variety of genome engineering approaches. Our reagents, based on transcription activator-like effector nucleases (TALENs) and the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system, are systematized for fast, modular cloning and accommodate diverse regulatory sequences to drive reagent expression. Vectors are optimized to create either single or multiple gene knockouts and large chromosomal deletions. Moreover, integration of geminivirus-based vectors enables precise gene editing through homologous recombination. Regulation of transcription is also possible. A Web-based tool streamlines vector selection and construction. One advantage of our platform is the use of the Csy-type (CRISPR system yersinia) ribonuclease 4 (Csy4) and tRNA processing enzymes to simultaneously express multiple guide RNAs (gRNAs). For example, we demonstrate targeted deletions in up to six genes by expressing 12 gRNAs from a single transcript. Csy4 and tRNA expression systems are almost twice as effective in inducing mutations as gRNAs expressed from individual RNA polymerase III promoters. Mutagenesis can be further enhanced 2.5-fold by incorporating the Trex2 exonuclease. Finally, we demonstrate that Cas9 nickases induce gene targeting at frequencies comparable to native Cas9 when they are delivered on geminivirus replicons. The reagents have been successfully validated in tomato (Solanum lycopersicum), tobacco (Nicotiana tabacum), Medicago truncatula, wheat (Triticum aestivum), and barley (Hordeum vulgare).

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

A Multipurpose Toolkit to Enable Advanced Genome Engineering in Plants

et al. 2017

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“…For example, SpCas9 could be driven by a nodule-specific promoter to effectively edit genes in Lotus japonicus by hairy root transformation [61]. In order to place the donor DNA and the CRISPR-Cas9 system in a context where HR is favored over NHEJ for DSB repair, the expression of the CRISPR elements can also be triggered or increased in the germlines by using cell-specific promoters [62,63]. Inducing genome editing in this type of cells also presents the advantage of limiting the generation of chimeric plants and ensuring a good inheritance of the targeted modifications.…”

Section: Nuclease Promotersmentioning

confidence: 99%

Towards mastering CRISPR-induced gene knock-in in plants: Survey of key features and focus on the model Physcomitrella patens

Collonnier

Guyon‐Debast

Maclot

et al. 2017

Methods

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a b s t r a c tBeyond its predominant role in human and animal therapy, the CRISPR-Cas9 system has also become an essential tool for plant research and plant breeding. Agronomic applications rely on the mastery of gene inactivation and gene modification. However, if the knock-out of genes by non-homologous end-joining (NHEJ)-mediated repair of the targeted double-strand breaks (DSBs) induced by the CRISPR-Cas9 system is rather well mastered, the knock-in of genes by homology-driven repair or end-joining remains difficult to perform efficiently in higher plants. In this review, we describe the different approaches that can be tested to improve the efficiency of CRISPR-induced gene modification in plants, which include the use of optimal transformation and regeneration protocols, the design of appropriate guide RNAs and donor templates and the choice of nucleases and means of delivery. We also present what can be done to orient DNA repair pathways in the target cells, and we show how the moss Physcomitrella patens can be used as a model plant to better understand what DNA repair mechanisms are involved, and how this knowledge could eventually be used to define more performant strategies of CRISPR-induced gene knock-in.

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“…Эта проблема была решена применением тка неспецифичных промоторов, обеспечивающих высокий уровень экспрессии Cas9 в зародышевых клетках (рис. 2) (Hyun et al, 2015;Wang et al, 2015;Yan et al, 2015;. Использование таких промоторов позволяет по лучать отредактированные растения в первом поколении трансгенных растений.…”

Section: стабильная трансформация растительных клеток компонентами сиunclassified

Making complex things simpler: modern tools to edit the plant genome

Zlobin¹,

Ternovoi²,

Grebenkina³

et al. 2017

Vestn. VOGiS

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There are several technologies for plant genome editing, of which the most simple and universal is CRISPR/ Cas. Currently, this technology is widely used for gene knockout, deleting genome fragments and inserting exogenous sequences in the plant genome. For each of these applications, many different types of genetic tools have been developed that are used by various research groups to solve specific problems. The CRISPR/Cas technology for plant genome editing is at an early stage of optimization, which is reflected by the ongoing search for the most effective, simple and flexible techniques. As a result, experimental work has to be preceded by a rather long and laborious process of selecting a genetic tool that will be optimal for a specific experimental task. In our review we describe the main variants of the CRISPR/Cas technology used to edit a plant genome. We classify them in terms of experimental tasks solved, major components and technology performance. In the first half of the review a detailed description of two major components of CRISPR/Cas technology -nuclease and guide RNAis given, the effect of structural features of these elements on editing efficiency is analyzed. Experimental data on the relationship between editing efficiency and nucleotide sequence of guide RNA are generalized. We also give the characteristic for different variants of nucleases used for plant genome editing and discuss their benefits for different experimental purposes. In the second half of the review various strategies for expression of CRISPR/Cas elements in plant cells, in particular, advantages and disadvantages of stable transformation and transient expression, are discussed. The effect of various regulatory elements of genes encoding nuclease and guide RNA on editing efficiency is described. Special emphasis is placed on the techniques of increasing targeted gene replacement efficiency.Key words: CRISPR/Cas; genome editing; plant genetic engineering; sgRNA.Существует несколько технологий редактирования генома рас-тений, из которых наиболее простой и универсальной является CRISPR/Cas. В настоящее время эта технология активно использу-ется для нокаутирования генов, делеций участков генома и встра-ивания экзогенных последовательностей в растительный геном. Для каждого из этих приложений разработано множество вариан-тов генетического инструментария, которые использовались раз-личными исследовательскими группами для решения конкретных задач. Технология CRISPR/Cas применительно к редактированию генома растений находится на начальном этапе оптимизации, вы-ражающейся в поиске наиболее эффективных, простых и универ-сальных методик. Вследствие этого экспериментальная работа должна предваряться достаточно длительным и трудоемким вы -бором варианта генетического инструментария, оптимального для решения конкретной экспериментальной задачи. В данном обзо-ре мы охарактеризовали разработанные на сегодняшний день основные варианты генетического инструментария технологии CRISPR/ Cas для редактирования генома растений с точки зрения решаем...

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High-Efficiency Genome Editing in Arabidopsis Using YAO Promoter-Driven CRISPR/Cas9 System

Abstract: We are grateful to Dr. Jiankang Zhu for providing us with the vector 35S-Cas9-SK. No conflict of interest declared.

Cited by 355 publications

References 14 publications

A Multipurpose Toolkit to Enable Advanced Genome Engineering in Plants

A Multipurpose Toolkit to Enable Advanced Genome Engineering in Plants

Towards mastering CRISPR-induced gene knock-in in plants: Survey of key features and focus on the model Physcomitrella patens

Making complex things simpler: modern tools to edit the plant genome

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