Exogenous DNA uptake by bovine spermatozoa does not induce DNA fragmentation

Feitosa, Weber Beringui; Mendes, Clayton Quirino; Milazzotto, Marcella Pécora; Rocha, A.M.; Martins, Lucas Carvalho; Simões, Rafael Plana; Paula-Lopes, F. F.; Visintin, José Antônio; Assumpção, M. E. O. A.

doi:10.1016/j.theriogenology.2010.03.016

Cited by 24 publications

(15 citation statements)

References 29 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…We used qRT‐PCR to reliably assess the number of exogenous DNA molecules internalized by sperm. This approach, which is the best technique available for the quantification of nucleic acids, has been successfully validated to quantify the uptake of foreign DNA into the spermatozoa of different animal species (Hoelker et al, 2007; Feitosa et al, 2010; Campos et al, 2011). We first established calibration curves for exogenous DNA content in samples, using the plasmid vector pcDNA 3.1/V5‐His and the β‐ actin gene as an internal control.…”

Section: Resultsmentioning

confidence: 99%

Introduction

Klump¹,

Steiger²

2013

Intl J Eating Disorders

View full text Add to dashboard Cite

Background Airborne enzymes behave as potent respiratory allergens. Till date, allergic disorders caused by genetically engineered enzymes widely used in the industry, have not been reported. Results and Conclusions We describe a worker employed in the detergent industry who developed asthma and rhinitis from IgE‐mediated sensitization to the thermostable endo‐alpha‐amylase Termamyl® and to the protease Savinase®. This is the first report showing that Termamyl® elicits allergic respiratory disorders in humans. Am. J. Ind. Med. 56:378–380, 2013. © 2012 Wiley Periodicals, Inc.

show abstract

Section: Resultsmentioning

confidence: 99%

Introduction

Klump¹,

Steiger²

2013

Intl J Eating Disorders

View full text Add to dashboard Cite

show abstract

“…Prolonged incubation before fertilization may increase the concentration of exogenous DNA associated with spermatozoa, but it may negatively affect sperm viability. Consequently, fertilization rates and the efficiency of exogenous DNA uptake was compromised, thereby reducing the rate of formation of viable transgenic embryos [53], [54].…”

Section: Discussionmentioning

confidence: 99%

First study of sperm mediated gene transfer in Egyptian river buffalo

Hassanane

Makawy

Helalia

et al. 2017

Journal of Genetic Engineering and Biotechnology

View full text Add to dashboard Cite

The present study was carried out to find the best treatments for enhancing the ration of insertion of a desired gene construct (pEGFP-N1) onto the sperm of buffalo as the first step for the production of transgenic buffalo using sperm mediated gene transfer (SMGT). The tested conditions were plasmid DNA concentration, sperm concentration, transfecting agent concentration: Dimethyle sulphoxide (DMSO) and time of transfection. The study proved that the best conditions for producing transgenic embryos were incubation sperm solution its concentration is 107/ml sperm with 3% DMSO: with 20 µg/ml from the linarized DNA, for 15 min at 4 °C are the best conditions to produce transgenic buffalo embryo using sperm mediated gene transfer.

show abstract

“…However, the theory has not been well documented in different laboratories across the globe (Eghbalsaied et al, 2015). The SMGT method, if proved to be reproducible, would be the simplest, costeffective, most rapid and mass gene transfer method to produce transgenic farm animals (Hoelker et al, 2007;Feitosa et al, 2010;Katebi et al, 2016). The SMGT strategy with the use of sex-sorted semen can also be an attractive approach to the generation of genetically-transformed male and female embryos/progeny derived from in vitro or in vivo fertilization (Zaniboni et al, 2016).…”

Section: Introductionmentioning

confidence: 99%

Expression of fluorescent reporter protein was not obtained in ovine embryos produced through in vitro fertilization-sperm mediated gene transfer (IVF-SMGT)

Hasanzadeh

Zandi

Sanjabi

et al. 2016

Small Ruminant Research

View full text Add to dashboard Cite

Sperm mediated gene transfer (SMGT) has been reported to be a powerful tool for producing transgenic animals on a mass scale using spermatozoa as vectors for exogenous DNA. In this study the possibility of using in vitro fertilization (IVF)-SMGT to produce transgenic ovine embryos was investigated for the first time. For this purpose, sperm were obtained from the epididymal testicle areas of 4 rams and different concentration of DNA (0.4, 0.8 and 1.6 μg) and TurboFect (0.25, 0.5, 1 and 2 μg) were used for sperm transfection (1×10 6). Enhanced-GFP-expressing vector pEGFP-N1 was used as the carrier. In order to evaluate the performance of transgenic sperm, in vitro fertilization technology was used. After the preparing oocytes received from the ovaries of slaughterhouse origin, oocytes with more than three layers of granulosa and uniform cytoplasm were selected and matured in TCM-199 medium containing 10% fetal calf serum, follicle stimulating hormone (FSH) (5μg/ml), β-17 estradiol (1μg/ml) and sodium pyruvate 0.81 (Mm). Bracket and Oliphant's (BO) medium and modified Charles Rosenkrans medium with amino acids (mCR2aa) were used for in vitro fertilization and culture, respectively. Results showed that transfected sperm with different concentrations of DNA and TurboFect carrier were unable to transfer the GFP gene to in vitro matured oocytes as the GFP gene was not expressed in neither zygotes nor morula stage embryos. Further optimization for SMGT improvement such as different transfection reagent and method like electroporation, using antioxidants in transfection medium to overcome the apoptosis and also separation of transfected sperm from untransfected ones before insemination have been suggested. Research highlights Transfected sperm with different concentrations of DNA and TurboFect carrier were unable to transfer the GFP gene to in vitro matured ovine oocytes. By increasing the amount of DNA, the expression of GFP gene was improved in fibroblast cells. The mean number of zygote and morula stage embryos significantly decreased, by increasing the amount of TurboFect.

show abstract

Exogenous DNA uptake by bovine spermatozoa does not induce DNA fragmentation

Cited by 24 publications

References 29 publications

Introduction

Introduction

First study of sperm mediated gene transfer in Egyptian river buffalo

Expression of fluorescent reporter protein was not obtained in ovine embryos produced through in vitro fertilization-sperm mediated gene transfer (IVF-SMGT)

Contact Info

Product

Resources

About