1. bookVolume 18 (2018): Issue 4 (October 2018)
Journal Details
License
Format
Journal
eISSN
2300-8733
First Published
25 Nov 2011
Publication timeframe
4 times per year
Languages
English
access type Open Access

Characterization of Three Generations of Transgenic Pigs Expressing the HLA-E Gene

Published Online: 02 Nov 2018
Volume & Issue: Volume 18 (2018) - Issue 4 (October 2018)
Page range: 919 - 935
Received: 08 Mar 2018
Accepted: 13 Jul 2018
Journal Details
License
Format
Journal
eISSN
2300-8733
First Published
25 Nov 2011
Publication timeframe
4 times per year
Languages
English
Abstract

The use of pigs as a source of organs and tissues for xenotransplantation can overcome the growing shortage of human donors. Human NK cells play an important role in the cell-mediated rejection of pig-to-human xenografts. In this paper we report the generation and extensive characterization of three generations of transgenic pigs with HLA-E gene encoding the antigen which can inhibit the human NK cell-mediated response. The gene construct pHLAE-GFPBsd containing the human gene encoding the human leukocyte antigen under the promoter of the EF-1α elongation factor ensuring systemic expression was introduced by microinjection into a pronucleus of the fertilized porcine oocyte. PCR analysis revealed and FISH analysis confirmed that the pHLAE-GFPBsd gene construct was present in the genome of the founder female pig. As a result of inter-breeding, an additional 7 transgenic animals were obtained (one individual from F1 generation and six individuals from F2 generation). The transgene expression was shown by RT-PCR and flow cytometry. Real Time PCR analysis estimated the approximate number of transgene copies at 16–34. Karyotype analysis did not show any changes in the structure or the number of chromosomes. The expression level of the transgene was stable in the next generation of genetically modified pigs. An NK cell-mediated cytotoxicity assay showed the increased viability of the transgenic cells in comparison with the wild-type, which confirmed the protective influence of HLA-E expression.

Keywords

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