Cell and Tissue BiologyCell and Tissue BiologyЦитология0041-37713034-6061The Russian Academy of Sciences67746610.31857/S0041377124050036DUVXRCArticlesСтатьиResearch ArticleDesign and selection of guides for CRISPR/Cas9-mediated knockout of the Kcnv2 gene in mouse cellsДизайн и подбор гидов для CRISPR/Cas9-опосредованного нокаута гена Kcnv2 в мышиных клеткахAntonovaE. N.АнтоноваЕ. Н.
Russian Federation
antonova.en@genlab.llcSorokaA. B.СорокаА. Б.
Russian Federation
antonova.en@genlab.llcMityaevaO. N.МитяеваО. Н.
Russian Federation
antonova.en@genlab.llcVolchkovP. Yu.ВолчковП. Ю.
Russian Federation
antonova.en@genlab.llcMoscow Institute of Physics and Technology (National Research University)Московский физико-технический институт (национальный исследовательский университет)Lomonosov Moscow State UniversityМосковский государственный университет имени М.В. ЛомоносоваFederal Research Center for Innovator and Emerging Biomedical and Pharmaceutical TechnologiesФедеральный исследовательский центр оригинальных и перспективных биомедицинских и фармацевтических технологий15112024665-642043720032025Copyright ©; 2024, Russian Academy of SciencesCopyright ©; 2024, Российская академия наук2024Russian Academy of SciencesРоссийская академия наукhttps://vietnamjournal.ru/0041-3771/article/view/677466

Mutations in the human KCNV2 gene cause a rare hereditary disease — cone dystrophy with supernormal rod response (CDSRR), characterized by progressive vision loss and impaired color discrimination. The KCNV2 gene encodes the Kv8.2 subunit of a potassium channel that is critical for the normal function of retinal photoreceptors. Gene therapy offers a promising treatment approach for this condition. To test the efficacy of gene therapy, an appropriate experimental disease model, such as a knockout mouse model, is required. This study focused on selecting optimal guide RNAs for knocking out the Kcnv2 gene using the CRISPR/Cas9 system and testing their efficiency in a mouse cell line. The selected guide RNAs can be utilized to generate a Kcnv2-/- mouse model.

Мутации в гене KCNV2 человека вызывают редкое наследственное заболевание сетчатки — дистрофию колбочек с повышенной активностью палочек (CDSRR), которая характеризуется прогрессирующей потерей зрения и нарушением цветоразличения. Ген KCNV2 кодирует субъединицу калиевого канала Kv8.2, играющую важную роль в нормальной работе фоторецепторов сетчатки. Поскольку в настоящее время этиотропной терапии для CDSRR не существует, генотерапия представляется перспективным методом лечения. Для проверки эффективности генотерапевтических подходов необходимо создание адекватной экспериментальной модели — нокаутной мышиной линии по гену Kcnv2. В нашем исследовании мы сосредоточились на подборе оптимальных гидовых РНК для нокаута данного гена с использованием системы CRISPR/Cas9 и их проверке на мышиной клеточной линии. Результаты экспериментов показали успешное создание одной маленькой 40 нуклеотидов (нт)) и нескольких больших делеций (1100–1400 нт) в гене Kcnv2. Эти данные подтверждают эффективность выбранных гидов и создают основу для разработки нокаутной мышиной модели, которая позволит изучать патофизиологию заболевания и провести тестирование потенциальных терапевтических методов.

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