Transfection into Mouse/Rat Cultured Embryos by Electroporation

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Transfection into Mouse/Rat Cultured Embryos by Electroporation

APPLICATIONS

Electroporation for mammalian embryos in the whole embryo culture system

Procedures

  1. ProcedureProcedure1: Pre-culture embryos in the whole embryo culture system for 1.5-2 hours prior to electroporation.
  2. Place the embryo in a Petri dish with Tyrode’s solution.
  3. Inject 0.1-0.5 μl of plasmid DNA into the brain ventricle with a fine capillary.
  4. Apply square pulses using the electroporator (CUY21) and electrodes (chamber-type and forceps-type electrodes are available). 70V, 50 msec at 1-second interval, five pulses are applied for E10.5 mouse embryos.
  5. Culture the electroporated embryos for 24-48 hours in the whole embryo culture system.Transfection of a fluorescent protein-expression vector into the developing rat cortex.

Transfection of a fluorescent protein-expression vector into the developing rat cortex.

(A) EGFP-expression vector was electroporated into E11.5 rat telencephalon. The electroporated embryo was cultured in the whole embryo culture system (WEC).

(B) 24 hours after electroporation, EGFP-expression was specifically detected at the dorsal part of the telencephalon.

Tel: telencephalon;
Di: diencephalon

Transfection of fluorescent protein-expression vectors into the rat spinal cord.

(A) EGFP-expression vector was transfected into the spinal cord of E12.5 rat embryos by electroporation. The electroporated embryos were cultured for 24 hours in the whole embryo culture system (WEC).

(B-E) Time-lapse analysis of neuroepithelial cells in the slice culture system. Histon-EGFP- and DsRed2-expression vectors were co-electroporated into the E12.0 rat spinal cord. The electroporated embryo was cultured for 24 hours in the WEC, then the spinal cord was sliced and time-lapse recording was performed.
(B-E) Cell nuclei (B, green) and cytoplasms (C, magenta) of neuroepithelial cells in the slice are simultaneously labeled by the electroporation
(E). (D) A DIC image.
hb: hindbrain; sc: spinal cord; fl: forelimb; DIC: differential interference contrast.

Masanori Takahashi, Tadashi Nomura, Noriko Osumi, Department of Developmental Neurobiology, Tohoku University Graduate School of Medicine
*Differentiation, Volume 70, Issue 4-5, Pages 155-162, June 2002

PUBLICATIONS

    Autotaxin/lysophospholipase D-mediated lysophosphatidic acid signaling is required to form distinctive large lysosomes in the visceral endoderm cells of the mouse yolk sac.

    Koike S, Keino-Masu K, Ohto T, Sugiyama F, Takahashi S, Masu M.

    J Biol Chem. 2009 Nov 27;284(48):33561-70.

      FGF8 signaling patterns the telencephalic midline by regulating putative key factors of midline development.

      Okada T, Okumura Y, Motoyama J, Ogawa M.

      Dev Biol. 2008 Aug 1;320(1):92-101.

        Isolation and Characterization of Vasohibin-2 as a Homologue of VEGF-Inducible Endothelium-Derived Angiogenesis Inhibitor Vasohibin

        Shibuya T, Watanabe K, Yamashita H, Shimizu K, Miyashita H, Abe M, Moriya T, Ohta H, Sonoda H, Shimosegawa T, Tabayashi K, Sato Y.

        Arterioscler Thromb Vasc Biol. 2006 May;26(5):1051-7.

          Electroporation-mediated gene transfer system applied to cultured CNS neurons

          Kawabata I, Umeda T, Yamamoto K, Okabe S.

          euroreport . 2004 Apr 29;15(6):971-5.

            Misrouting of mitral cell progenitors in the Pax6/small eye rat telencephalon

            Nomura T, Osumi N.

            Development . 2004 Feb;131(4):787-96.

              Manipulating gene expressions by electroporation in the developing brain of mammalian embryos

              Takahashi M, Sato K, Nomura T, Osumi N.

              Differentiation. 2002 Jun;70(4-5):155-62.

                Pax6 regulates specification of ventral neurone subtypes in the hindbrain by establishing progenitor domains

                Takahashi M, Osumi N.

                Development . 2002 Mar;129(6):1327-38.

                  Gene Transfer into Cultured Mammalian Embryos by Electroporation

                  Osumi N, Inoue T.

                  Methods . 2001 May;24(1):35-42.

                  Electroporation

                  Drug Delivery and Transfection

                  Electro Cell Fusion

                  Fluorescent Staining

                  Single-Cell/Micro-Particle Transfer

                  Cell Freezing

                  Mechanical Vibration