Exogenous DNA uptake by South American catfish (Rhamdia quelen) spermatozoa after seminal plasma removal

Campos, Vinícius Farias; Amaral, Marta; Seixas, Fabiana Kömmling; Pouey, Juvencio Luis Fernandez Osório; Selau, Lisiane Priscila Roldão; Dellagostin, Odir A.; Deschamps, João Carlos; Collares, Tiago

doi:10.1016/j.anireprosci.2011.05.004

Cited by 12 publications

(6 citation statements)

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“…The fi rst report that exogenous DNA could be introduced into sperm was made by Brackett et al (1971). Several studies in distinct species have reported the generation of transgenic animals using spermatozoa as vectors to carrier foreign DNA to the ova (Lu et al, 2002;Webster et al, 2005;Shen et al, 2006;Hoelker et al, 2007, Campos et al, 2011a, Campos et al, 2011b, Campos et al, 2011c. One approach of sperm-mediated gene transfer (SMGT) is the direct introduction of foreign DNA into testes, so-called testis-mediated gene transfer (TMGT), which allows for natural mating and mass gene transfer.…”

Section: Introductionmentioning

confidence: 99%

Testis-mediated gene transfer in mice: comparison of transfection reagents regarding transgene transmission and testicular damage

et al. 2011

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Testis-mediated gene transfer (TMGT) has been used as in vivo gene transfer technology to introduce foreign DNA directly into testes, allowing mass gene transfer to offspring via mating. In this study, we used plasmid DNA (pEGFP-N1) mixed with dimethylsulfoxide (DMSO), N,N-dimethylacetamide (DMA) or liposome (Lipofectin) in an attempt to improve TMGT. Males receiving consecutive DNA complex injections were mated to normal females to obtain F0 progeny. In vivo evaluation of EGFP expression, RT-PCR and PCR were used to detect the expression and the presence of exogenous DNA in the progeny. We also evaluated possible testicular damage by histological procedures. PCR and RT-PCR analyses revealed that liposome and DMSO increased the rate of TMGT. Histological analyses demonstrated that repeated (4 times) injections of DNA complexes can affect spermatogenesis. DMSO was the most deleterious among the reagents tested. In this study, we detected the presence of transgene in the progeny, and its expression in blood cells. Consecutive injections of DNA complexes were associated with impaired spermatogenesis, suggesting requirement of optimal conditions for DNA delivery through TMGT.

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Section: Introductionmentioning

confidence: 99%

Testis-mediated gene transfer in mice: comparison of transfection reagents regarding transgene transmission and testicular damage

et al. 2011

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“…Several approaches have been utilized in order to improve DNA uptake. Methods such as electroporation, lipofection, DNA/DMSO complexes and restriction-enzyme-mediated integration (REMI) have been used; however the frequency of transgenic offspring remains low [6,8,21,43]. Conversely, the more recent use of nanotechnology in the context of SMGT has new possibilities for the efficient delivery of exogenous DNA into sperm.…”

Section: Nanotubes For Dna Transfectionmentioning

confidence: 99%

The Use of Nanostructures for DNA Transfection

Campos

Yurgel

Seixas

et al. 2012

Carbon Nanostructures

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The interaction between nanostructured materials and living systems is of fundamental and practical interest and will determine the biocompatibility, potential utilities and applications of novel nanomaterials in biological settings. The pursuit of new types of molecular transporters is an active area of research, due to the high impermeability of cell membranes and other biological barriers to foreign substances and the need for intercellular delivery of molecules via cell-penetrating transporter for drug, gene or protein therapeutics. Here, is described the novel nanostructurebased transfection systems. The transfection uses of nanopolymers, nanoparticles and nanotubes are the main focus of this review. In addition are described the technique called NanoSMGT that uses nanostructures for DNA transfection in sperm cells that could be used for transgenic animal generation or human gene therapy.

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“…Most studies only investigate the gene transfer efficiency, being found to be between 20–30% using conventional electroporation [ 12 ], without evaluating the transfection rate in fish sperm. Some studies have investigated the negative effects of seminal plasma DNase on sperm DNA uptake, however, no reliable data are available for optimization of electroporation for fish spermatozoa [ 15 , 16 ].…”

Section: Introductionmentioning

confidence: 99%

“…Zebrafish is also being considered as a possible animal model to study COVID-19 [ 18 ]. Nevertheless, laborious microinjection is still the standard method to generate transgenic zebrafish [ 15 ]. Until now, few studies been performed for zebrafish transgenesis using SMGT with controversial results [ 9 , 13 , 19 ].…”

Section: Introductionmentioning

confidence: 99%

Efficiency and cell viability implications using tip type electroporation in zebrafish sperm cells

et al. 2020

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Sperm-mediated gene transfer (SMGT) has a potential use for zebrafish transgenesis. However, transfection into fish sperm cells still needs to be improved. The objective was to demonstrate the feasibility of tip type electroporation in zebrafish sperm, showing a protocol that provide high transfection efficiency, with minimal side-effects. Sperm was transfected with a Cy3-labelled DNA using tip type electroporation with voltages ranging from 500 to 1500 V. Sperm kinetics parameters were assessed using Computer Assisted Semen Analysis (CASA) and cell integrity, reactive oxygen species (ROS), mitochondrial functionality and transfection rate were evaluated by flow cytometry. The transfection rates were positively affected by tip type electroporation, reaching 64.9% ± 3.6 in the lowest voltage used (500 V) and 86.6% ± 1.9 in the highest (1500 V). The percentage of overall motile sperm in the electrotransfected samples was found to decrease with increasing field strength (P < 0.05). Increase in the sperm damaged plasma membrane was observed with increasing field strength (P < 0.05). ROS and sperm mitochondrial functionality did not present a negative response after the electroporation (P > 0.05). Overall results indicate that tip type electroporation enhances the internalization of exogenous DNA into zebrafish sperm cells with minimal harmful effects to sperm cells.

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Exogenous DNA uptake by South American catfish (Rhamdia quelen) spermatozoa after seminal plasma removal

Cited by 12 publications

References 20 publications

Testis-mediated gene transfer in mice: comparison of transfection reagents regarding transgene transmission and testicular damage

Testis-mediated gene transfer in mice: comparison of transfection reagents regarding transgene transmission and testicular damage

The Use of Nanostructures for DNA Transfection

Efficiency and cell viability implications using tip type electroporation in zebrafish sperm cells

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