Efficient expression of transgenes in adult zebrafish by electroporation

Rambabu, K. Murali; Rao, Shu-Mei; Rao, Nalam Madhusudhana

doi:10.1186/1472-6750-5-29

Cited by 33 publications

(10 citation statements)

References 18 publications

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“…Nowadays, the EP method is used in cancer research (Canatella and Prausnitz 2001;Jamieson et al 1989;Nishi et al 1996;Yang and Sun 1995), gene packaged with lipids, peptides and viruses transfection (Coulberson et al 2003;Tryfona and Bustard 2006), and drug delivery (Canatella and Prausnitz 2001). In addition, the EP method is successful in transgenic treatments in mice (Lavitrano et al 1989;Marti et al 2004;Neumann et al 1982;Nishi et al 1996), silkworm (Shamila and Mathavan 1998), sea urchin (Larochelle and Epel 1991), cattle (Gagne et al 1991), chick (Nakamura andFunahashi 2001), abalone (Powers et al 1995), and fish (Rambabu et al 2005;Connaughton et al 2004). …”

Section: Introductionmentioning

confidence: 95%

An electroporation microchip system for the transfection of zebrafish embryos using quantum dots and GFP genes for evaluation

Huang¹,

Lin²,

Su³

et al. 2007

Biomed Microdevices

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This study focuses on the design and experimental verification of an electroporation (EP) microchip system for the transfection of zebrafish (Danio rerio). For generating suitable pulses, a circuit is used to provide voltages between 0 and 700 V, with nearly 0-3,500 V/cm electric field. In addition, a proposed EP microchip, designed in a modular fashion, is fabricated using micro electromechanical system (MEMS) technology to allow for rapid and convenient replacement of each component. A numerical simulation is carried out to analyze the uniformity and strength of the EP electric fields generated in the microchip. Trypan blue dye, water-soluble quantum dots (MUA-QDs) and genes coding for green fluorescence protein (pEGFP-N1 plasmids) were employed to verify the successful delivery and transfection of zebrafish embryos. The experimental results show that the optimum delivery rate of trypan blue dyes and MUA-QDs were respectively up to 62 and 36% by using the proposed EP system. The successfully transfected embryos with the pEGFP-N1 plasmid used exhibit green fluorescence in the zebrafish embryos. The approach in the transfection of zebrafish embryos will provide many potential usages for cellular imaging areas, gene therapy research and medical applications.

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Section: Introductionmentioning

confidence: 95%

An electroporation microchip system for the transfection of zebrafish embryos using quantum dots and GFP genes for evaluation

Huang¹,

Lin²,

Su³

et al. 2007

Biomed Microdevices

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show abstract

“…Electroporation is a well-characterized method for molecule delivery into embryos of many species (Lurquin 1997;Swartz et al 2001), although microinjection is more common for zebrafish embryos. As might be expected, compound trans-membrane permeability is highest for cells nearest to the electrodes (Rambabu et al 2005). The amplitude and frequency of the applied potential, as well as its overall shape and total duration are usually empirically determined for each cell type; a range exists wherein membrane pores reseal themselves after delivery (Rambabu et al 2005).…”

Section: Device Theorymentioning

confidence: 99%

“…As might be expected, compound trans-membrane permeability is highest for cells nearest to the electrodes (Rambabu et al 2005). The amplitude and frequency of the applied potential, as well as its overall shape and total duration are usually empirically determined for each cell type; a range exists wherein membrane pores reseal themselves after delivery (Rambabu et al 2005). Excessive field strength or long durations cause irreversible damage to the cells and the organism (Lurquin 1997).…”

Section: Device Theorymentioning

confidence: 99%

“…) in suspension are introduced between two opposite electrodes. A high electric field, typically from 1 to 10 kV/cm, is applied to the electrodes to deliver the molecules into the cells (Huang et al 2007;Rambabu et al 2005;Lurquin 1997). Electroporation is a well-characterized method for molecule delivery into embryos of many species (Lurquin 1997;Swartz et al 2001), although microinjection is more common for zebrafish embryos.…”

Section: Device Theorymentioning

confidence: 99%

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Patterned delivery and expression of gene constructs into zebrafish embryos using microfabricated interfaces

Bansal

Lenhart

Kim

et al. 2009

Biomed Microdevices

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We demonstrate a method which uses simple microfabrication and microfluidics to produce custom, shaped electroporators for the patterned delivery of foreign molecules into developing embryos. We show how these electroporators can be used to 'draw' two-dimensional patterns of tracer molecules, DNA and mRNA into the yolk and cells of zebrafish embryos (Danio rerio) at different stages of development. We demonstrate the successful delivery of patterns of Trypan Blue (normal dye), Texas Red (fluorescent dye), GFP-expressing DNA plasmids and GFP expressing mRNA constructs into both chorionated and dechorionated embryos. Both DNA and mRNA were expressed in the desired patterns subsequent to delivery. Square pulses of 10-20 V (0.20-0.40 kV/cm), 50-100 ms width were sufficient to create transient pores and introduce compounds from the late blastula period (3 hpf) to early pharyngula period (24 hpf) embryos. Using 24 hpf dechorionated embryos, we achieved a high survival of 91.3% and 89%, and a delivery efficiency of 38% and 50% for GFP-DNA and GFP-mRNA respectively. Lastly, we demonstrate the simultaneous delivery of different compounds into the developing embryo.

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“…The major hurdles for plasmid-mediated gene therapy are active exploration [6,7]. However, some degrees of studies in zebrafish gene therapy are currently under way [8].…”

Section: Introductionmentioning

confidence: 99%

Expression of Green Fluorescence Protein (GFP) in Zebrafish Muscle through Injection: A Gene Therapy Model

et al. 2008

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Expression of the target gene is important for gene therapy. Presently, localized transgenesis is used for gene therapy which can be achieved by a target gene expression. Here, we have reported the plasmid mediated gene therapy to zebrafish model. For this purpose, we have chosen green fluorescent protein (GFP) as a target gene because the expression can be detected easily. GFP was inserted in a plasmid vector, pQE30 to develop the vector pQE30GFP. The plasmid pQE30GFP was constructed form plasmid, pQE30 and pEGFPC2. pQE30GFP injected directly in one group of fish into the muscle where luciferase expression was noted. In another group, after injection electroporation was performed where we have also noted luciferase expression; but, electroporation cause muscle injury to the zebrafish. In our case, the expression was very strong at the site of injection in first group in compare to electroporation group and in both the cases expression was stable more than two weeks.

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Efficient expression of transgenes in adult zebrafish by electroporation

Cited by 33 publications

References 18 publications

An electroporation microchip system for the transfection of zebrafish embryos using quantum dots and GFP genes for evaluation

An electroporation microchip system for the transfection of zebrafish embryos using quantum dots and GFP genes for evaluation

Patterned delivery and expression of gene constructs into zebrafish embryos using microfabricated interfaces

Expression of Green Fluorescence Protein (GFP) in Zebrafish Muscle through Injection: A Gene Therapy Model

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