Efficiency of transgene expression in bovine cells varies according to cell type and gene transfer method

Authors

  • Alinne G. Curcio State University of Northern Rio de Janeiro Darcy Ribeiro
  • Fabiana F. Bressan University of São Paulo
  • Carla S. Paes De Carvalho State University of Northern Rio de Janeiro Darcy Ribeiro
  • Célia R. Quirino State University of Northern Rio de Janeiro Darcy Ribeiro
  • Flavio V. Meirelles University of São Paulo
  • Angelo J. B. Dias State University of Northern Rio de Janeiro Darcy Ribeiro

DOI:

https://doi.org/10.17533/udea.rccp.v32n1a04

Keywords:

cloning, epigenetics, lipofection, lentiviral transduction, nuclear reprogramming

Abstract

Background: Production of transgenic animals is still a low-efficiency biotechnology, and simple alternatives should be used to improve the rate of transgenic bovine production by nuclear transfer. One such alternative is selecting the appropriate donor cell type and transfection method. Objective: To investigate the effect of cell type (fetal or adult fibroblasts, and cumulus cells), and gene transfer method (lipofection and lentiviral transduction) on the incorporation, expression, and fluorescence intensity of transgene on bovine cells analyzed by flow cytometry. Methods: Fetal fibroblasts (FF), adult fibroblasts (AF), and cumulus cells (CC) were transfected using lipofection, or transduced using lentiviral particles produced with Green Fluorescent Protein (GFP) expressing plasmids, and analyzed by flow cytometry. Results: Lentiviral transduction resulted in higher transgene expression rates for all cell types (FF: 88.8 ± 0.98; AF: 91.6 ± 2.96; CC: 60.7% ± 14.7) compared to lipofection (FF: 17.8 ± 2.82; AF: 10.66 ± 0.65; CC: 3.9% ± 1.97). Cumulus cells showed lower transgene expression rates than the other cell types. Regarding fluorescence intensity, there was no significant difference between lipofection and lentiviral transduction; in both treatments, higher fluorescence intensity was obtained when adult cells were used instead of fetal cells. Conclusion: Gene transfer efficiency varies according to cell type, and gene transfer method, with lentiviral transduction achieving higher transgene expression rate, and adult fibroblasts showing better transgene expression.

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Author Biographies

Alinne G. Curcio, State University of Northern Rio de Janeiro Darcy Ribeiro

VM, MSc., Laboratory of Animal Reproduction and Genetic Improvement, Center for Agricultural Sciences and Technologies, State University of Northern Rio de Janeiro Darcy Ribeiro (UENF), Campos dos Goytacazes, Rio de Janeiro.

Fabiana F. Bressan, University of São Paulo

VM, PhD., Department of Veterinary Medicine, Faculty of Zootechnics and Food Engineering, University of São Paulo (USP), Pirassununga, São Paulo, Brazil.

Carla S. Paes De Carvalho, State University of Northern Rio de Janeiro Darcy Ribeiro

VM, PhD., Laboratory of Animal Reproduction and Genetic Improvement, Center for Agricultural Sciences and Technologies, State University of Northern Rio de Janeiro Darcy Ribeiro (UENF), Campos dos Goytacazes, Rio de Janeiro, Brazil.

 

Célia R. Quirino, State University of Northern Rio de Janeiro Darcy Ribeiro

Agr. Eng., PhD., Laboratory of Animal Reproduction and Genetic Improvement, Center for Agricultural Sciences and Technologies, State University of Northern Rio de Janeiro Darcy Ribeiro (UENF), Campos dos Goytacazes, Rio de Janeiro, Brazil.

Flavio V. Meirelles, University of São Paulo

VM, PhD., Department of Veterinary Medicine, Faculty of Zootechnics and Food Engineering, University of São Paulo (USP), Pirassununga, São Paulo, Brazil.

Angelo J. B. Dias, State University of Northern Rio de Janeiro Darcy Ribeiro

VM, PhD., Laboratory of Animal Reproduction and Genetic Improvement, Center for Agricultural Sciences and Technologies, State University of Northern Rio de Janeiro Darcy Ribeiro (UENF), Campos dos Goytacazes, Rio de Janeiro.

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Published

2019-03-27

How to Cite

Curcio, A. G., Bressan, F. F., Paes De Carvalho, C. S., Quirino, C. R., Meirelles, F. V., & Dias, A. J. B. (2019). Efficiency of transgene expression in bovine cells varies according to cell type and gene transfer method. Revista Colombiana De Ciencias Pecuarias, 32(1), 34–42. https://doi.org/10.17533/udea.rccp.v32n1a04

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