Transfection into Chick Embryos in Ovo/Others by Electroporation
APPLICATIONS
Misexpression of the gene of interest by in ovo electroporation
Knock-down by transfection of shRNA expression vector by in ovo electroporation
Harukazu Nakamura and Tatsuya Katahira, Graduate School of Life Sciences, and Institute of Development, Aging and Cancer, Tohoku University
*Mechanisms of Development, Volume 121, Issue 9, Pages 1137-1143, September 2004
*Development Growth & Differentiation, Volume 45, Issue 4, Pages 361-367, August 2003
Electroporation for early chick embryos using New's culture (gastrula)
- Schema illustrating the system.
- Hamburger and Hamilton (HH) stage 4 chick embryo with DNA solution injected to the prospective neural plate region. DNA solution was colored with a FastGreen dye.hn: Hensen’s node, ps: primitive streak, ao: area opaca, ap: area pellucida
- Schema illustrating the location of DNA solution between the vitelline membrane and the ectoderm.
Experimental Procedure
- Dissect an embryo using a paper ring (dried filter paper) out of the egg.
- Rinse gently to remove excess yolk, and place the embryo onto the platform of the cathode chamber.
- Inject DNA solution into the space between the ectoderm and vitelline membrane using a glass pipette (for introduction to the ectoderm derivatives).
- Adjust the target site right onto the cathode, and set the anode above it. Set the electrode distance at 5mm.
- Deliver electric pulses (10V, pulse on 50ms, pulse of 100ms, 5 pulses) using a CUY21 electroporator.
- Place the specimen on an agarose-albumin plate, and incubate at 39C
Transgene expression after the electroporation
Gene transfection for tissues other than the CNS
Kenji Shimamura, Division of Morphogenesis, Institute of Molecular Embryology and Genetics, Kumamoto University
PUBLICATIONS
- In_Ovo
- Anterior_primitive_streak
Mechanical Coupling Coordinates the Co-elongation of Axial and Paraxial Tissues in Avian Embryos
Xiong F, Ma W, Bénazéraf B, Mahadevan L, Pourquié O.
Dev Cell. 2020 Nov 9;55(3):354-366.e5.
- In_Ovo
- Ciliary ganglion(CG)
Analysis of Neuro-Neuronal Synapses Using Embryonic Chick Ciliary Ganglion via Single-Axon Tracing, Electrophysiology, and Optogenetic Techniques
Egawa R, Yawo H.
Curr Protoc Neurosci. 2019 Apr;87(1):e64.
- In_Ovo
- Neural_plate
- Neural_tube
Method for electroporation for the early chick embryo
Hatakeyama J, Shimamura K.
Dev Growth Differ. 2008 Aug;50(6):449-52.
- Ex_Ovo
- Neural_plate
- Neural_tube
Apical Accumulation of Rho in the Neural Plate Is Important for Neural Plate Cell Shape Change and Neural Tube Formation
Kinoshita N, Sasai N, Misaki K, Yonemura S.
Mol Biol Cell. 2008 May;19(5):2289-99.
- Ex_Ovo
- Anterior_primitive_streak
Dual mode of paraxial mesoderm formation during chick gastrulation
Iimura T, Yang X, Weijer CJ, Pourquié O.
PNAS, Volume 104, Number 8, Pages 2744-2749, 20 February 2007
- In_Ovo
- Neural_tube
Gain- and loss-of-function in chick embryos by electroporation
Nakamura H, Katahira T, Sato T, Watanabe Y, Funahashi J.
Mech Dev. 2004 Sep;121(9):1137-43.
- In_Ovo
- Neural_tube
- Brain
RGM and its receptor neogenin regulate neuronal survival
Matsunaga E, Tauszig-Delamasure S, Monnier PP, Mueller BK, Strittmatter SM, Mehlen P, Chédotal A.
Nat Cell Biol. 2004 Aug;6(8):749-55.
- Ex_Ovo
- Somites
A primer on using in ovo electroporation to analyze gene function
Krull CE.
Dev Dyn. 2004 Mar;229(3):433-9.
- In_Ovo
- Primitive_cardiac_tube
- Ex_Ovo
- Cardiac_explants
Dual roles of Sema6D in cardiac morphogenesis through region-specific association of its receptor, Plexin-A1, with off-track and vascular endothelial growth factor receptor type 2
Toyofuku T, Zhang H, Kumanogoh A, Takegahara N, Suto F, Kamei J, Aoki K, Yabuki M, Hori M, Fujisawa H, Kikutani H.
Genes Dev . 2004 Feb 15;18(4):435-47.
- In_Ovo
- Neural_fold
- Neural_tube
- Ex_Ovo
- Neural_tube_explants
Multiple roles of Sox2, an HMG-box transcription factor in avian neural crest development
Wakamatsu Y, Endo Y, Osumi N, Weston JA.
Dev Dyn. 2004 Jan;229(1):74-86.
- In_Ovo
- Neural_tube
Gene silencing in chick embryos with a vector-based small interfering RNA system
Katahira T, Nakamura H.
Dev Growth Differ. 2003 Aug;45(4):361-7.
- In_Ovo
- prospective_esophago_tracheobronchial_region
Tbx4-Fgf10 system controls lung bud formation during chicken embryonic development
Sakiyama J, Yamagishi A, Kuroiwa A.
Development. 2003 Apr;130(7):1225-34.
- In_Ovo
- Neural_tube
Inductive signal and tissue responsiveness defining the tectum and the cerebellum
Sato T, Araki I, Nakamura H.
Development . 2001 Jul;128(13):2461-9.
Electroporation
■ Cell Cultures
- Primary Cell Cultures
- Stem Cells
- Organoids
- Cell Lines
- Cells in Adherence
■ In Vivo Mice/Rats
- Zygotes In Vitro (TAKE method)
- Zygotes In Oviduct (i-GONAD method)
- Embryos In Utero
- Ex Utero Embryos In Vitro
- Brain
- Retina / Cornea / Spinal Cord / Sciatic Nerve
- Lung / Spleen / Liver / Stomach/ Kidney / Intestine
- Pancreas / Islets of langerhans
- Testis / Ovary / Prostate / Gonad / Uterus
- Muscle / Skin / Joint / Cartilage / Tumor / Others
■ In Vivo Other Animals
- Bovine/Porcine/Other Animal Zygotes
- Hamster Zygotes in Oviduct (i-GONAD method)
- Monkey Skin
- Chicken (In Ovo・Others)
- Zebrafish & Other Fishes
- Insects・Others
■ Plant Cells & Algae
- Plant Cells
- Algae
■ Exosomes
- Exosomes
■ Bacteria, Yeast, Fungi
- E. coli and bacteria
- Yeast and Fungi
- Bacteria, Yeast, Fungi (NEPA Porator)
Drug Delivery and Transfection
■ Ultrasound Transfection and Drug Delivery (Sonoporation/Fus)
- Brain
- Liver/Skin/Other Applications
- Heart
- Cell Culture
- Lung
- Muscle
Electro Cell Fusion
■ Hybridoma Production
- Monoclonal antibodies, etc
■ Oocyte Activation
- Electrical stimulation before/after Intracytoplasmic sperm injection (ICSI)
■ Somatic cell nuclear transfer (SCNT)・ Oocytes Nuclear Transfer
- Animal cloning
■ Tetraploid Embryos Production
- 2 Cell Embryos (Tetraploid)
■ Other Applications
- Liposome・Protoplast・Yeast, etc.
Fluorescent Staining
■ Autofluorescence Quenching
- Mammalian Tissue Sections
- Fish Tissue Sections
- Amphibia tissue sections
- Avian Tissue Sections
- Plant tissue sections
Single-Cell/Micro-Particle Transfer
■ Micro targets
- Animal cells
■ Micro liquid
- Plant cells
Cell Freezing
■ Cell Therapy
- Stem cells, primary cells, etc.
■ Animal Husbandry
- Sperm, embryos, tissue, etc.