## The negative effects of exogenous DNA binding on porcine spermatozoa are caused by removal of seminal fluid

2008
##### Authors
Hakimov, Hatam
Ruiz, Arturo
Friendship, Robert M.
Buhr, Mary
Golovan, Serguei P.
Journal article
##### Published in
Theriogenology ; 70 (2008), 8. - pp. 1288-1296. - ISSN 0093-691X
##### Abstract
Sperm-mediated gene transfer (SMGT) might become the most efficient and cost effective technique to generate transgenic animals, which will significantly increase their application in biomedical research and in commercial production. Despite some successes, the technique has remained controversial for almost 20 years and despite number of studies the reasons for poor reproducibility of this promising technology has not been understood. We suggest that the reason for poor reproducibility is the presence of natural defences against exogenous DNA invasion acting in spermatozoa or in embryo. Based on previous reports we have investigated the effect of foreign DNA binding on spermatozoa by monitoring motility, viability and genomic DNA damage. Evaluation of DNA binding in sperm collected from 16 boars demonstrated that 28 45% of the added pEGFP plasmid was bound to spermatozoa with 9 32% being internalized in sperm nucleus. In agreement with previous reports, our results demonstrated that the pEGFP-treated sperm show an average a 2-fold decrease in motility ( p < 0.05), 5-fold decrease in progressive motility ( p < 0.05), and 1.4-fold increase in number of sperm with highly damaged DNA ( p < 0.05) as detected by Comet assay. In contrast with previous reports, we demonstrate that all such changes were associated with the removal of seminal plasma during the washing step and not with foreign DNA binding per se. We suggest that poor reproducibility of SMGT most likely result from selection against DNA-loaded sperm at later stages of fertilization.
##### Subject (DDC)
570 Biosciences, Biology
##### Keywords
Sperm-mediated gene transfer,Comet assay,Spermatozoa,DNA damage,Seminal plasma
##### Cite This
ISO 690KANG, Ji Hyoun, Hatam HAKIMOV, Arturo RUIZ, Robert M. FRIENDSHIP, Mary BUHR, Serguei P. GOLOVAN, 2008. The negative effects of exogenous DNA binding on porcine spermatozoa are caused by removal of seminal fluid. In: Theriogenology. 70(8), pp. 1288-1296. ISSN 0093-691X. Available under: doi: 10.1016/j.theriogenology.2008.06.011
BibTex
@article{Kang2008negat-6707,
year={2008},
doi={10.1016/j.theriogenology.2008.06.011},
title={The negative effects of exogenous DNA binding on porcine spermatozoa are caused by removal of seminal fluid},
number={8},
volume={70},
issn={0093-691X},
journal={Theriogenology},
pages={1288--1296},
author={Kang, Ji Hyoun and Hakimov, Hatam and Ruiz, Arturo and Friendship, Robert M. and Buhr, Mary and Golovan, Serguei P.}
}

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<dcterms:abstract xml:lang="eng">Sperm-mediated gene transfer (SMGT) might become the most efficient and cost effective technique to generate transgenic animals, which will significantly increase their application in biomedical research and in commercial production. Despite some successes, the technique has remained controversial for almost 20 years and despite number of studies the reasons for poor reproducibility of this promising technology has not been understood. We suggest that the reason for poor reproducibility is the presence of natural defences against exogenous DNA invasion acting in spermatozoa or in embryo. Based on previous reports we have investigated the effect of foreign DNA binding on spermatozoa by monitoring motility, viability and genomic DNA damage. Evaluation of DNA binding in sperm collected from 16 boars demonstrated that 28 45% of the added pEGFP plasmid was bound to spermatozoa with 9 32% being internalized in sperm nucleus. In agreement with previous reports, our results demonstrated that the pEGFP-treated sperm show an average a 2-fold decrease in motility ( p &lt; 0.05), 5-fold decrease in progressive motility ( p &lt; 0.05), and 1.4-fold increase in number of sperm with highly damaged DNA ( p &lt; 0.05) as detected by Comet assay. In contrast with previous reports, we demonstrate that all such changes were associated with the removal of seminal plasma during the washing step and not with foreign DNA binding per se. We suggest that poor reproducibility of SMGT most likely result from selection against DNA-loaded sperm at later stages of fertilization.</dcterms:abstract>
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