Protocol for assessment of the efficiency of CRISPR/Cas RNP delivery to different types of target cells

<h4>Background</h4> Delivery of CRISPR/Cas RNPs to target cells still remains the biggest bottleneck to genome editing. Many efforts are made to develop efficient CRISPR/Cas RNP delivery methods that will not affect viability of target cell dramatically. Popular current methods and proto...

Descripción completa

Guardado en:

Detalles Bibliográficos
Autores principales:	Marina A. Tyumentseva, Aleksandr I. Tyumentsev, Vasiliy G. Akimkin
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/29be6d3838f34a0492e637a62af0d06e
Etiquetas:	Agregar Etiqueta Sin Etiquetas, Sea el primero en etiquetar este registro!

id	oai:doaj.org-article:29be6d3838f34a0492e637a62af0d06e
record_format	dspace
spelling	oai:doaj.org-article:29be6d3838f34a0492e637a62af0d06e2021-11-18T06:34:24ZProtocol for assessment of the efficiency of CRISPR/Cas RNP delivery to different types of target cells1932-6203https://doaj.org/article/29be6d3838f34a0492e637a62af0d06e2021-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC8577758/?tool=EBIhttps://doaj.org/toc/1932-6203<h4>Background</h4> Delivery of CRISPR/Cas RNPs to target cells still remains the biggest bottleneck to genome editing. Many efforts are made to develop efficient CRISPR/Cas RNP delivery methods that will not affect viability of target cell dramatically. Popular current methods and protocols of CRISPR/Cas RNP delivery include lipofection and electroporation, transduction by osmocytosis and reversible permeabilization and erythrocyte-based methods. <h4>Methods</h4> In this study we will assess the efficiency and optimize current CRISPR/Cas RNP delivery protocols to target cells. We will conduct our work using molecular cloning, protein expression and purification, cell culture, flow cytometry (immunocytochemistry) and cellular imaging techniques. <h4>Discussion</h4> This will be the first extensive comparative study of popular current methods and protocols of CRISPR/Cas RNP delivery to human cell lines and primary cells. All protocols will be optimized and characterized using the following criteria i) protein delivery and genome editing efficacy; ii) viability of target cells after delivery (post-transduction recovery); iii) scalability of delivery process; iv) cost-effectiveness of the delivery process and v) intellectual property rights. Some methods will be considered ‘research-use only’, others will be recommended for scaling and application in the development of cell-based therapies.Marina A. TyumentsevaAleksandr I. TyumentsevVasiliy G. AkimkinPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 16, Iss 11 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Marina A. Tyumentseva Aleksandr I. Tyumentsev Vasiliy G. Akimkin Protocol for assessment of the efficiency of CRISPR/Cas RNP delivery to different types of target cells
description	<h4>Background</h4> Delivery of CRISPR/Cas RNPs to target cells still remains the biggest bottleneck to genome editing. Many efforts are made to develop efficient CRISPR/Cas RNP delivery methods that will not affect viability of target cell dramatically. Popular current methods and protocols of CRISPR/Cas RNP delivery include lipofection and electroporation, transduction by osmocytosis and reversible permeabilization and erythrocyte-based methods. <h4>Methods</h4> In this study we will assess the efficiency and optimize current CRISPR/Cas RNP delivery protocols to target cells. We will conduct our work using molecular cloning, protein expression and purification, cell culture, flow cytometry (immunocytochemistry) and cellular imaging techniques. <h4>Discussion</h4> This will be the first extensive comparative study of popular current methods and protocols of CRISPR/Cas RNP delivery to human cell lines and primary cells. All protocols will be optimized and characterized using the following criteria i) protein delivery and genome editing efficacy; ii) viability of target cells after delivery (post-transduction recovery); iii) scalability of delivery process; iv) cost-effectiveness of the delivery process and v) intellectual property rights. Some methods will be considered ‘research-use only’, others will be recommended for scaling and application in the development of cell-based therapies.
format	article
author	Marina A. Tyumentseva Aleksandr I. Tyumentsev Vasiliy G. Akimkin
author_facet	Marina A. Tyumentseva Aleksandr I. Tyumentsev Vasiliy G. Akimkin
author_sort	Marina A. Tyumentseva
title	Protocol for assessment of the efficiency of CRISPR/Cas RNP delivery to different types of target cells
title_short	Protocol for assessment of the efficiency of CRISPR/Cas RNP delivery to different types of target cells
title_full	Protocol for assessment of the efficiency of CRISPR/Cas RNP delivery to different types of target cells
title_fullStr	Protocol for assessment of the efficiency of CRISPR/Cas RNP delivery to different types of target cells
title_full_unstemmed	Protocol for assessment of the efficiency of CRISPR/Cas RNP delivery to different types of target cells
title_sort	protocol for assessment of the efficiency of crispr/cas rnp delivery to different types of target cells
publisher	Public Library of Science (PLoS)
publishDate	2021
url	https://doaj.org/article/29be6d3838f34a0492e637a62af0d06e
work_keys_str_mv	AT marinaatyumentseva protocolforassessmentoftheefficiencyofcrisprcasrnpdeliverytodifferenttypesoftargetcells AT aleksandrityumentsev protocolforassessmentoftheefficiencyofcrisprcasrnpdeliverytodifferenttypesoftargetcells AT vasiliygakimkin protocolforassessmentoftheefficiencyofcrisprcasrnpdeliverytodifferenttypesoftargetcells
_version_	1718424467782238208

Protocol for assessment of the efficiency of CRISPR/Cas RNP delivery to different types of target cells

Ejemplares similares