Транзиентная экспрессия генов в растениях ‒ эффективная экспериментальная платформа для функциональной геномики

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Большой массив данных об экспрессии генов растений, накопленный благодаря сравнительным исследованиям, направляет усилия исследователей для изучения тонких механизмов влияния целевых генов и, как следствие, на разработку относительно простых и одновременно эффективных подходов, позволяющих понять физиологическую роль белковых продуктов гена. Многочисленные исследования убедительно продемонстрировали эффективность стратегии транзиентной экспрессии для характеристики функций генов растений. Цели обзора: (i) рассмотреть преимущества и ограничения различных растительных систем и методов временной экспрессии, используемых для выяснения роли генных продуктов; (ii) обобщить текущие данные об использовании подходов к временной экспрессии для понимания тонких механизмов, лежащих в основе функции генов; (iii) описать достижения в области эффективной временной экспрессии генов растений. В обзоре обсуждаются основные и критические этапы каждого из методов транзиентной экспрессии генов у растений, области их применения, а также основные результаты, полученные с использованием растительных объектов и их вклад в наши знания о тонких механизмах функций генов, лежащих в основе роста и развития растений, включая выяснение механизмов, регулирующих сложные метаболические пути.

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作者简介

И. Голденкова-Павлова

Федеральное государственное бюджетное научное учреждение Институт физиологии растений им. К.А. Тимирязева Российской академии наук

编辑信件的主要联系方式.
Email: irengold58@gmail.com
俄罗斯联邦, Москва

О. Павленко

Федеральное государственное бюджетное научное учреждение Институт физиологии растений им. К.А. Тимирязева Российской академии наук

Email: irengold58@gmail.com
俄罗斯联邦, Москва

И. Демьянчук

Федеральное государственное бюджетное научное учреждение Институт физиологии растений им. К.А. Тимирязева Российской академии наук

Email: irengold58@gmail.com
俄罗斯联邦, Москва

В. Фридман

Федеральное государственное бюджетное научное учреждение Институт физиологии растений им. К.А. Тимирязева Российской академии наук

Email: irengold58@gmail.com
俄罗斯联邦, Москва

А. Тюрин

Федеральное государственное бюджетное научное учреждение Институт физиологии растений им. К.А. Тимирязева Российской академии наук

Email: irengold58@gmail.com
俄罗斯联邦, Москва

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2. Fig. 1. Plant systems used for transient expression and their delivery to plant cells: (a) intact plants, isolated organs or specialized plant tissues, the main methods of delivery are agroinfiltration, viral particles, bioballistics; (b) callus, suspension cultures of plant cells, the main methods of delivery are bioballistics; (c) – protoplasts of plant cells, the main delivery methods are electroporation.

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3. Fig. 2. Scheme of T-DNA gene constructs for various aspects of studying gene products in a plant cell: (a) – a general scheme of the T-DNA region, including a promoter and terminator (Promoter, Terminator – mandatory components); enhancer (Enhancer, cloned when testing the functional activity of sequences); leader signal (Leader, cloned when studying the localization of the gene product in a plant cell); target gene and/or reporter gene (Goi and/or Reporter – target gene is cloned when studying the functional role of the gene product of the target gene; transcriptional-translational fusion of the target gene with the reporter gene is used in studying the localization of the gene product in a plant cell, as well as in protein-protein interaction studies; the reporter gene is used in testing the functional activity of sequences); (b) – Agrobacteria are transformed by gene constructs, which are used in transient expression to (c) study the locale

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4. Fig. 3. Scheme of protein-protein interactions using bimolecular fluorescent complementation analysis (BiFC) and a three–way complementation system: (a, b) - the BiFC system includes two interacting proteins, each of which has a transcriptional-translational fusion with one of the protein domains of the fluorescent protein (a), due to the joint folding of the fluorescent protein in the case of interaction of target proteins, which leads to the restoration of the fluorophore (d). (b, c, e) – the tripartite complementation system includes two GFP fragments, each of which has a transcriptional-translational fusion with one of the studied partner proteins, and an additional system with additional GFP fragments (b), during protein-protein interaction (c) additional GFP fragments interact with GFP fragments that are fused with partner proteins, which leads to the restoration of a full-size GFP that fluoresces (e). The principle of using more

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5. Fig. 4. The main directions of research using transient gene expression in plants. The delivery of gene constructs to plant systems using various methodological approaches expands the experimental capabilities of researchers in the field of functional genomics of plants. BBB – protein-protein interaction.

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