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In Vitro Cell Dev Biol Anim
2011 Oct 01;479:640-52. doi: 10.1007/s11626-011-9451-2.
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Optimization of gene delivery methods in Xenopus laevis kidney (A6) and Chinese hamster ovary (CHO) cell lines for heterologous expression of Xenopus inner ear genes.
Ramirez-Gordillo D
,
Trujillo-Provencio C
,
Knight VB
,
Serrano EE
.
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The Xenopus inner ear provides a useful model for studies of hearing and balance because it shares features with the mammalian inner ear, and because amphibians are capable of regenerating damaged mechanosensory hair cells. The structure and function of many proteins necessary for inner ear function have yet to be elucidated and require methods for analysis. To this end, we seek to characterize Xenopus inner ear genes outside of the animal model through heterologous expression in cell lines. As part of this effort, we aimed to optimize physical (electroporation), chemical (lipid-mediated; Lipofectamine™ 2000, Metafectene® Pro), and biological (viral-mediated; BacMam virus Cellular Lights™ Tubulin-RFP) gene delivery methods in amphibian (Xenopus; A6) cells and mammalian (Chinese hamster ovary (CHO)) cells. We successfully introduced the commercially available pEGFP-N3, pmCherry-N1, pEYFP-Tubulin, and Cellular Lights™ Tubulin-RFP fluorescent constructs to cells and evaluated their transfection or transduction efficiencies using the three gene delivery methods. In addition, we analyzed the transfection efficiency of a novel construct synthesized in our laboratory by cloning the Xenopus inner ear calcium-activated potassium channel β1 subunit, then subcloning the subunit into the pmCherry-N1 vector. Every gene delivery method was significantly more effective in CHO cells. Although results for the A6 cell line were not statistically significant, both cell lines illustrate a trend towards more efficient gene delivery using viral-mediated methods; however the cost of viral transduction is also much higher. Our findings demonstrate the need to improve gene delivery methods for amphibian cells and underscore the necessity for a greater understanding of amphibian cell biology.
Altmann,
Insect cells as hosts for the expression of recombinant glycoproteins.
1999, Pubmed
Altmann,
Insect cells as hosts for the expression of recombinant glycoproteins.
1999,
Pubmed
Balls,
Amphibian cells in vitro. II. Effects of variations in medium osmolarity on a permanent cells line isolated from Xenopus.
1973,
Pubmed
,
Xenbase
Banerjee,
Heterologous expression of the serotonin 5-HT1A receptor in neural and non-neural cell lines.
1993,
Pubmed
Boyer,
The need for water quality criteria for frogs.
1995,
Pubmed
Butel,
Demonstration of infectious DNA in transformed cells. III. Correlation of detection of infectious DNA-protein complexes with persistence of virus in simian adenovirus SA7-induced tumor cells.
1975,
Pubmed
Chesneau,
Transgenesis procedures in Xenopus.
2008,
Pubmed
,
Xenbase
Condreay,
Transient and stable gene expression in mammalian cells transduced with a recombinant baculovirus vector.
1999,
Pubmed
Desai,
Production of heterologous proteins in plants: strategies for optimal expression.
2010,
Pubmed
Duncan,
Variation in large-conductance, calcium-activated potassium channels from hair cells along the chicken basilar papilla.
2003,
Pubmed
Díaz,
Quantity, bundle types, and distribution of hair cells in the sacculus of Xenopus laevis during development.
1995,
Pubmed
,
Xenbase
Faletti,
sgk: an essential convergence point for peptide and steroid hormone regulation of ENaC-mediated Na+ transport.
2002,
Pubmed
,
Xenbase
Fettiplace,
Mechanisms of hair cell tuning.
1999,
Pubmed
Hellsten,
The genome of the Western clawed frog Xenopus tropicalis.
2010,
Pubmed
,
Xenbase
Ibey,
Plasma membrane permeabilization by 60- and 600-ns electric pulses is determined by the absorbed dose.
2009,
Pubmed
Ichigi,
Dome formation and tubule morphogenesis by Xenopus kidney A6 cell cultures exposed to microgravity simulated with a 3D-clinostat and to hypergravity.
2001,
Pubmed
,
Xenbase
Kim,
Systematic investigation of polyamidoamine dendrimers surface-modified with poly(ethylene glycol) for drug delivery applications: synthesis, characterization, and evaluation of cytotoxicity.
2008,
Pubmed
Knight,
Tissue and species differences in the application of quantum dots as probes for biomolecular targets in the inner ear and kidney.
2006,
Pubmed
,
Xenbase
Liu,
Identification and authentication of animal cell culture by polymerase chain reaction amplification and DNA sequencing.
2003,
Pubmed
Long,
Functional entry of baculovirus into insect and mammalian cells is dependent on clathrin-mediated endocytosis.
2006,
Pubmed
Lopez-Novoa,
New insights into the mechanism of aminoglycoside nephrotoxicity: an integrative point of view.
2011,
Pubmed
Matsuki,
Low voltage pulses can induce apoptosis.
2008,
Pubmed
Mies,
Epithelial Na+ channel stimulation by n-3 fatty acids requires proximity to a membrane-bound A-kinase-anchoring protein complexed with protein kinase A and phosphodiesterase.
2007,
Pubmed
,
Xenbase
Nakahara,
Effects of exposure of CHO-K1 cells to a 10-T static magnetic field.
2002,
Pubmed
Oberholtzer,
Frequency tuning of cochlear hair cells by differential splicing of BK channel transcripts.
1999,
Pubmed
Orio,
New disguises for an old channel: MaxiK channel beta-subunits.
2002,
Pubmed
Pyott,
Cochlear function in mice lacking the BK channel alpha, beta1, or beta4 subunits.
2007,
Pubmed
Ramanathan,
A molecular mechanism for electrical tuning of cochlear hair cells.
1999,
Pubmed
Scolnick,
Isolation of infectious xenotropic mouse type C virus by transfection of a heterologous cell with DNA from a transformed mouse cell.
1975,
Pubmed
Serrano,
Identification of genes expressed in the Xenopus inner ear.
2001,
Pubmed
,
Xenbase
Shin,
Xenopus TRPN1 (NOMPC) localizes to microtubule-based cilia in epithelial cells, including inner-ear hair cells.
2005,
Pubmed
,
Xenbase
Smith,
Production of human beta interferon in insect cells infected with a baculovirus expression vector.
1983,
Pubmed
Tanaka,
Proliferation and differentiation of Xenopus A6 cells under hypergravity as revealed by time-lapse imaging.
2003,
Pubmed
,
Xenbase
Terhag,
Cave Canalem: how endogenous ion channels may interfere with heterologous expression in Xenopus oocytes.
2010,
Pubmed
,
Xenbase
Trujillo-Provencio,
RNA isolation from Xenopus inner ear sensory endorgans for transcriptional profiling and molecular cloning.
2009,
Pubmed
,
Xenbase
Varela-Ramírez,
Detection of transcripts for delayed rectifier potassium channels in the Xenopus laevis inner ear.
1998,
Pubmed
,
Xenbase
