Lyse-Reseal Erythrocytes for Transfection of Plasmodium falciparum
| Autor: | Deepak Kumar, Gokulapriya Govindarajalu, Zeba Rizvi, Puran Singh Sijwali |
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| Rok vydání: | 2019 |
| Předmět: |
0301 basic medicine
Parasitic infection animal structures Erythrocytes Lysis Cost effectiveness Science viruses Plasmodium falciparum Transfection Article 03 medical and health sciences chemistry.chemical_compound 0302 clinical medicine Plasmid parasitic diseases Lysis buffer Humans Malaria Falciparum Multidisciplinary biology Chemistry Electroporation fungi Gene Transfer Techniques DNA biology.organism_classification Cell biology 030104 developmental biology embryonic structures Medicine 030217 neurology & neurosurgery Plasmids |
| Zdroj: | Scientific Reports Scientific Reports, Vol 9, Iss 1, Pp 1-8 (2019) |
| ISSN: | 2045-2322 |
| DOI: | 10.1038/s41598-019-56513-9 |
| Popis: | 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. |
| Databáze: | OpenAIRE |
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