References

vavilov

Вавиловский журнал генетики и селекции

Vavilov Journal of Genetics and Breeding

2500-3259

Institute of Cytology and Genetics of Siberian Branch of the RAS

10.18699/VJ17.228

vavilov-909

Research Article

Перспективные направления

Promising trends

Сделать сложное проще: современный инструментарий для редактирования генома растений

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

Злобин

Н. Е.

Zlobin

N. E.

Москва

Moscow

Терновой

В. В.

Ternovoy

V. V.

Москва

Moscow

Гребенкина

Н. А.

Grebenkina

N. A.

Москва

Moscow

Таранов

В. В.

Taranov

V. V.

Москва

Moscow

v.taranov@iab.ac.ru

Федеральное государственное бюджетное научное учреждение «Всероссийский научно-исследовательский институт сельскохозяйственной биотехнологии»РоссияAll-Russia Research Institute of Agricultural BiotechnologyRussian Federation

2017

24032017

211104111

2017

Злобин Н.Е., Терновой В.В., Гребенкина Н.А., Таранов В.В.

Zlobin N.E., Ternovoy V.V., Grebenkina N.A., Taranov V.V.

This work is licensed under a Creative Commons Attribution 4.0 License.

https://vavilov.elpub.ru/jour/article/view/909

Существует несколько технологий редактирования генома растений, из которых наиболее простой и универсальной является CRISPR/Cas. В настоящее время эта технология активно используется для нокаутирования генов, делеций участков генома и встраивания экзогенных последовательностей в растительный геном. Для каждого из этих приложений разработано множество вариантов 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, выражающейся в поиске наиболее эффективных, простых и универсальных методик. Вследствие этого экспериментальная работа должна предваряться достаточно длительным и трудоемким выбором варианта генетического инструментария, оптимального для решения конкретной экспериментальной задачи. В данном обзоре мы охарактеризовали разработанные на сегодняшний день основные варианты генетического инструментария технологии CRISPR/Cas для редактирования генома растений с точки зрения решаемых экспериментальных задач, составляющих компонентов и эффективности применения. В первой части подробно рассмотрены основные элементы технологии CRISPR/Cas – нуклеаза и направляющая РНК, проанализировано влияние структурных особенностей этих элементов на эффективность редактирования. Обобщены экспериментальные данные о взаимосвязи между эффективностью редактирования и нуклеотидной последовательностью направляющей РНК. Охарактеризованы различные варианты нуклеаз, использовавшиеся при редактировании геномов растений, обсуждаются преимущества этих вариантов для решения определенных экспериментальных задач. Вторая часть обзора посвящена различным стратегиям экспрессии элементов системы CRISPR/Cas в растительных клетках, в частности преимуществам и недостаткам использования стабильной трансформации и транзиентной экспрессии. Описывается влияние регуляторных элементов генов, кодирующих нуклеазу и направляющую РНК, на эффективность редактирования. Особый акцент сделан на способах повышения эффективности замещения целевых участков в гене растений на экзогенные последовательности ДНК.

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 RNA – is 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.

CRISPR/Casредактирование геномагенетическая инженерия растенийsgRNA

CRISPR/Casgenome editingplant genetic engineeringsgRNA

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The authors declare that there are no conflicts of interest present.