Lyse-Reseal Erythrocytes for Transfection of Plasmodium falciparum

Abstract Simple and efficient transfection methods for genetic manipulation of Plasmodium falciparum are desirable to identify, characterize and validate the genes with therapeutic potential and better understand parasite biology. Among the available transfection techniques for P. falciparum, electr...

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Autores principales:	Gokulapriya Govindarajalu, Zeba Rizvi, Deepak Kumar, Puran Singh Sijwali
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2019
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/2ad36a5cb9fe4cf584b58fdab4b364fe
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spelling	oai:doaj.org-article:2ad36a5cb9fe4cf584b58fdab4b364fe2021-12-02T13:35:03ZLyse-Reseal Erythrocytes for Transfection of Plasmodium falciparum10.1038/s41598-019-56513-92045-2322https://doaj.org/article/2ad36a5cb9fe4cf584b58fdab4b364fe2019-12-01T00:00:00Zhttps://doi.org/10.1038/s41598-019-56513-9https://doaj.org/toc/2045-2322Abstract Simple and efficient transfection methods for genetic manipulation of Plasmodium falciparum are desirable to identify, characterize and validate the genes with therapeutic potential and better understand parasite biology. Among the available transfection techniques for P. falciparum, electroporation-based methods, particularly electroporation of ring-infected RBCs is routinely used. Nonetheless, transfection of P. falciparum remains a resource-intensive procedure. Here, we report a simple and economic transfection method for P. falciparum, which is termed as the lyse-reseal erythrocytes for transfection (LyRET). It involved lysis of erythrocytes with a hypotonic RBC lysis buffer containing the desired plasmid DNA, followed by resealing by adding a high salt buffer. These DNA-encapsulated lyse-reseal erythrocytes were mixed with P. falciparum trophozoite/schizont stages and subjected to selection for the plasmid-encoded drug resistance. In parallel, transfections were also done by the methods utilizing electroporation of DNA into uninfected RBCs and parasite-infected RBCs. The LyRET method successfully transfected 3D7 and D10 strains with different plasmids in 63 of the 65 attempts, with success rate similar to transfection by electroporation of DNA into infected RBCs. The cost effectiveness and comparable efficiency of LyRET method makes it an alternative to the existing transfection methods for P. falciparum, particularly in resource-limited settings.Gokulapriya GovindarajaluZeba RizviDeepak KumarPuran Singh SijwaliNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 9, Iss 1, Pp 1-8 (2019)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Gokulapriya Govindarajalu Zeba Rizvi Deepak Kumar Puran Singh Sijwali Lyse-Reseal Erythrocytes for Transfection of Plasmodium falciparum
description	Abstract Simple and efficient transfection methods for genetic manipulation of Plasmodium falciparum are desirable to identify, characterize and validate the genes with therapeutic potential and better understand parasite biology. Among the available transfection techniques for P. falciparum, electroporation-based methods, particularly electroporation of ring-infected RBCs is routinely used. Nonetheless, transfection of P. falciparum remains a resource-intensive procedure. Here, we report a simple and economic transfection method for P. falciparum, which is termed as the lyse-reseal erythrocytes for transfection (LyRET). It involved lysis of erythrocytes with a hypotonic RBC lysis buffer containing the desired plasmid DNA, followed by resealing by adding a high salt buffer. These DNA-encapsulated lyse-reseal erythrocytes were mixed with P. falciparum trophozoite/schizont stages and subjected to selection for the plasmid-encoded drug resistance. In parallel, transfections were also done by the methods utilizing electroporation of DNA into uninfected RBCs and parasite-infected RBCs. The LyRET method successfully transfected 3D7 and D10 strains with different plasmids in 63 of the 65 attempts, with success rate similar to transfection by electroporation of DNA into infected RBCs. The cost effectiveness and comparable efficiency of LyRET method makes it an alternative to the existing transfection methods for P. falciparum, particularly in resource-limited settings.
format	article
author	Gokulapriya Govindarajalu Zeba Rizvi Deepak Kumar Puran Singh Sijwali
author_facet	Gokulapriya Govindarajalu Zeba Rizvi Deepak Kumar Puran Singh Sijwali
author_sort	Gokulapriya Govindarajalu
title	Lyse-Reseal Erythrocytes for Transfection of Plasmodium falciparum
title_short	Lyse-Reseal Erythrocytes for Transfection of Plasmodium falciparum
title_full	Lyse-Reseal Erythrocytes for Transfection of Plasmodium falciparum
title_fullStr	Lyse-Reseal Erythrocytes for Transfection of Plasmodium falciparum
title_full_unstemmed	Lyse-Reseal Erythrocytes for Transfection of Plasmodium falciparum
title_sort	lyse-reseal erythrocytes for transfection of plasmodium falciparum
publisher	Nature Portfolio
publishDate	2019
url	https://doaj.org/article/2ad36a5cb9fe4cf584b58fdab4b364fe
work_keys_str_mv	AT gokulapriyagovindarajalu lyseresealerythrocytesfortransfectionofplasmodiumfalciparum AT zebarizvi lyseresealerythrocytesfortransfectionofplasmodiumfalciparum AT deepakkumar lyseresealerythrocytesfortransfectionofplasmodiumfalciparum AT puransinghsijwali lyseresealerythrocytesfortransfectionofplasmodiumfalciparum
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Lyse-Reseal Erythrocytes for Transfection of Plasmodium falciparum

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