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In Utero Intraventriculaire injectie en elektroporatie van de E16 Rat embryo's

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Institute for Regeneration Medicine, University of California, San Francisco - UCSF

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Cite this Article: In Utero Intraventriculaire injectie en elektroporatie van de E16 Rat embryo's

Walantus, W., Elias, L., Kriegstein, A. In Utero Intraventricular Injection and Electroporation of E16 Rat Embryos. J. Vis. Exp. (6), e236, doi:10.3791/236 (2007).

Abstract: In Utero Intraventriculaire injectie en elektroporatie van de E16 Rat embryo's

In-utero in-vivo injectie en elektroporatie van de embryonale rat neocortex is een krachtig hulpmiddel voor het manipuleren van individuele voorouders langs de wanden van de laterale ventrikel. Deze techniek wordt nu op grote schaal gebruikt om de processen die betrokken zijn bij corticogenesis door over-uitdrukken of neerhalen genen en het observeren van de effecten op de cellulaire proliferatie, migratie en differentiatie te bestuderen. In vergelijking met traditionele knock-out strategieën, in-utero elektroporatie biedt een snelle manier om een ​​populatie van cellen te manipuleren tijdens een bepaalde temporele venster. In deze video protocol, schetsen we de experimentele methode voor het bereiden van ratten voor een operatie, waardoor de baarmoeder hoornen door laporatomy, het injecteren van DNA in de laterale ventrikels van het zich ontwikkelende embryo, electroporating DNA in de voorouders langs de laterale wand, en de zorg voor dieren, post- operatie. Ons laboratorium maakt gebruik van dit protocol voor de operaties op E15-E21 ratten, maar het wordt meestal uitgevoerd op E16 zoals in deze video.

Disclosures: In Utero Intraventriculaire injectie en elektroporatie van de E16 Rat embryo's

Ask the Author: In Utero Intraventriculaire injectie en elektroporatie van de E16 Rat embryo's

7 Comments

Very nice work. I do this routinely in Mouse but have little luck with Rat. I get poor survival of pups. If I do a sham experiment, I get fine survival with electroporation only but little survival if I inject only. I've used a variety of anesthetics. Any suggestions particular to Rat? Thank you much

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Posted by: AnonymousOctober 24, 2007, 2:50 PM

Try different methods of purifying the DNA.

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Posted by: AnonymousOctober 29, 2007, 5:19 PM

I am having poor survival as well. Any ideas about different methods of purification? I am currently using a Promega MIDI/Maxi kit. Have you noticed with purification methods are better?

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Posted by: Chris CunninghamOctober 18, 2010, 1:53 PM

hi barry. are you beveling your injection pipettes enough? if they are not sharp enough embryos will have high mortality. have a look at the fourth chapter of the video again.

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Posted by: Aaron K.October 31, 2007, 2:12 AM

Thanks, will try, but mice go just fine without problems except that B6 mice don't make the best of moms.

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Posted by: AnonymousOctober 31, 2007, 8:53 AM

Hi, I would appreciate if you could send me the protocol for mice ...Thanks a lot !
Martine
m.barkats@institut-myologie.org

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Posted by: Martine B.May 28, 2010, 11:40 AM

Where can we find the tools used to make the syrine injector tip?

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Posted by: DerekJuly 17, 2008, 2:59 PM

what are the medicine for motor neuron disease?

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Posted by: saraSeptember 8, 2009, 1:23 AM

From which company can I buy the electrods for the electroporation. The same as you have. Could you tell me their cat. # also?

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Posted by: Julie CardinNovember 10, 2009, 4:19 PM


BTX Harvard Apparatus Tweezertrodes, you can get them from Fisher and other vendors. -William

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Posted by: AnonymousNovember 10, 2009, 4:40 PM

I was not able to find the volume for these intraventricular infusions. Also, what is the maximum infusion volume for these kinds of procedures using newborn rats? Thanks!

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Posted by: mark b.August 7, 2010, 8:49 AM

I was not able to find the volume for these intraventricular infusions. Also, what is the maximum infusion volume for these kinds of procedures using newborn rats? Thanks!

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Posted by: mark b.August 7, 2010, 8:50 AM

Have you ever delivered DNA earlier into the lateral ventricles (E12.5 or E13)?

Thank you.

Shelley

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Posted by: Shelley RussekNovember 30, 2011, 11:22 PM

No. Sorry, There are some who do though. You can use ultrasound or IR videomicroscopy.

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Posted by: BarryDecember 1, 2011, 4:17 PM

Thank you...any one in particular that you know has a technique working or a paper published? Thanks!

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Posted by: ShelleyDecember 1, 2011, 4:56 PM

Thank you...any one in particular that you know has a technique working or a paper published? Thanks!

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Posted by: ShelleyDecember 1, 2011, 4:57 PM

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