Effect of custom-designed transfection buffer on delivery of genome modification components into primary cells of buffalo, cattle, goats, and sheep
The transfer of genome-modification components into farm animal cells is indispensable for the production of genome-modified and transgenic farm animals. Electroporation is a physical transfection method when appropriately used; this technique is safe, simple to use, affordable, and efficient in tr...
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Universidad del Zulia
2023-11-01
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| Series: | Revista Científica |
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| Online Access: | https://mail.produccioncientificaluz.org/index.php/cientifica/article/view/43486 |
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| author | Shreya Malhotra Priyanka Singh Aseem Tara Bosco Jose Devika Gautam Ram Prasad Gaurav Tripathi Sacchinandan De Naresh L. Selokar |
| author_facet | Shreya Malhotra Priyanka Singh Aseem Tara Bosco Jose Devika Gautam Ram Prasad Gaurav Tripathi Sacchinandan De Naresh L. Selokar |
| author_sort | Shreya Malhotra |
| collection | DOAJ |
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The transfer of genome-modification components into farm animal cells is indispensable for the production of genome-modified and transgenic farm animals. Electroporation is a physical transfection method when appropriately used; this technique is safe, simple to use, affordable, and efficient in transfecting cells from several lineages. Electroporation efficiency depends on various physical parameters, of which cell type is considered a major factor for transfection efficiency. Primary cells are generally less susceptible to transfection than other cell types due to their finite lifespan and limited expansion capacity. Previously, we custom-designed a transfection buffer to deliver exogenous genetic components into mammalian cells. In the present study, we examined the effect of the developed buffer on transfection rates and cell viability of primary somatic cells from buffalo, cattle, goats, and sheep. To achieve the aims of this study, t primary somatic cells from skin biopsies were established and were transfected with a Venus-expression vector (pAcGFPs-Venus). We noticed that transfection rates of pAcGFPs-Venus were 22.51%, 17.56%, 22.81%, and 16.16% for buffalo, cattle, goats, and sheep cells, respectively. We also noticed that cell viability and proliferation rates were better in the case of goats, sheep, and cattle cells; also, these cells have less vacuolation than buffalo cells. In addition, we also generated MSTN (myostatin) KO (Knockout) cell clones from these cell populations, in which the efficiency of single-cell clone generation was high for goats and sheep cells. In conclusion, our lab-made transfection buffer can be efficiently used to generate genome-edited or transgenic farm animals for agriculture, biomedical, and veterinary applications.
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| format | Article |
| id | doaj-art-b4377be46125482fac4515cf1b56f2cb |
| institution | Kabale University |
| issn | 0798-2259 2521-9715 |
| language | English |
| publishDate | 2023-11-01 |
| publisher | Universidad del Zulia |
| record_format | Article |
| series | Revista Científica |
| spelling | doaj-art-b4377be46125482fac4515cf1b56f2cb2025-02-06T15:37:07ZengUniversidad del ZuliaRevista Científica0798-22592521-97152023-11-0133Suplemento10.52973/rcfcv-wbc123Effect of custom-designed transfection buffer on delivery of genome modification components into primary cells of buffalo, cattle, goats, and sheepShreya Malhotra0Priyanka Singh1Aseem Tara2Bosco Jose3Devika Gautam4Ram Prasad5Gaurav Tripathi6Sacchinandan De7Naresh L. Selokar8Animal Biotechnology Division (ABTD), ICAR-National Dairy Research Institute, Karnal, Haryana, 132001, IndiaAnimal Biotechnology Division (ABTD), ICAR-National Dairy Research Institute, Karnal, Haryana, 132001, IndiaAnimal Biotechnology Division (ABTD), ICAR-National Dairy Research Institute, Karnal, Haryana, 132001, IndiaAnimal Biotechnology Division (ABTD), ICAR-National Dairy Research Institute, Karnal, Haryana, 132001, IndiaAnimal Biotechnology Division (ABTD), ICAR-National Dairy Research Institute, Karnal, Haryana, 132001, IndiaAnimal Biotechnology Division (ABTD), ICAR-National Dairy Research Institute, Karnal, Haryana, 132001, IndiaAnimal Biotechnology Division (ABTD), ICAR-National Dairy Research Institute, Karnal, Haryana, 132001, IndiaAnimal Biotechnology Division (ABTD), ICAR-National Dairy Research Institute, Karnal, Haryana, 132001, IndiaAnimal Biotechnology Division (ABTD), ICAR-National Dairy Research Institute, Karnal, Haryana, 132001, India The transfer of genome-modification components into farm animal cells is indispensable for the production of genome-modified and transgenic farm animals. Electroporation is a physical transfection method when appropriately used; this technique is safe, simple to use, affordable, and efficient in transfecting cells from several lineages. Electroporation efficiency depends on various physical parameters, of which cell type is considered a major factor for transfection efficiency. Primary cells are generally less susceptible to transfection than other cell types due to their finite lifespan and limited expansion capacity. Previously, we custom-designed a transfection buffer to deliver exogenous genetic components into mammalian cells. In the present study, we examined the effect of the developed buffer on transfection rates and cell viability of primary somatic cells from buffalo, cattle, goats, and sheep. To achieve the aims of this study, t primary somatic cells from skin biopsies were established and were transfected with a Venus-expression vector (pAcGFPs-Venus). We noticed that transfection rates of pAcGFPs-Venus were 22.51%, 17.56%, 22.81%, and 16.16% for buffalo, cattle, goats, and sheep cells, respectively. We also noticed that cell viability and proliferation rates were better in the case of goats, sheep, and cattle cells; also, these cells have less vacuolation than buffalo cells. In addition, we also generated MSTN (myostatin) KO (Knockout) cell clones from these cell populations, in which the efficiency of single-cell clone generation was high for goats and sheep cells. In conclusion, our lab-made transfection buffer can be efficiently used to generate genome-edited or transgenic farm animals for agriculture, biomedical, and veterinary applications. https://mail.produccioncientificaluz.org/index.php/cientifica/article/view/43486transfection buffergenome modificationCRISPR |
| spellingShingle | Shreya Malhotra Priyanka Singh Aseem Tara Bosco Jose Devika Gautam Ram Prasad Gaurav Tripathi Sacchinandan De Naresh L. Selokar Effect of custom-designed transfection buffer on delivery of genome modification components into primary cells of buffalo, cattle, goats, and sheep Revista Científica transfection buffer genome modification CRISPR |
| title | Effect of custom-designed transfection buffer on delivery of genome modification components into primary cells of buffalo, cattle, goats, and sheep |
| title_full | Effect of custom-designed transfection buffer on delivery of genome modification components into primary cells of buffalo, cattle, goats, and sheep |
| title_fullStr | Effect of custom-designed transfection buffer on delivery of genome modification components into primary cells of buffalo, cattle, goats, and sheep |
| title_full_unstemmed | Effect of custom-designed transfection buffer on delivery of genome modification components into primary cells of buffalo, cattle, goats, and sheep |
| title_short | Effect of custom-designed transfection buffer on delivery of genome modification components into primary cells of buffalo, cattle, goats, and sheep |
| title_sort | effect of custom designed transfection buffer on delivery of genome modification components into primary cells of buffalo cattle goats and sheep |
| topic | transfection buffer genome modification CRISPR |
| url | https://mail.produccioncientificaluz.org/index.php/cientifica/article/view/43486 |
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