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In Utero Intraventricular Injection and Electroporation of E16 Rat Embryos
William Walantus, Laura Elias, Arnold Kriegstein
Institute for Regeneration Medicine, University of California, San Francisco

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0:00 Title0

0:17 Introduction17

0:21 Subscription Lock21

1:06 Tools and Reagents66

2:17 Preparation of Microinjection Pipets137

6:23 Prepping Rat for Surgery383

10:02 Removing Uterus and Exposing Embryos602

12:12 In utero Injection and Electroporation732

14:11 Placing Embryos Back Inside Dam851

17:23 Conclusion1043

18:48 End Credits1128

In-utero in-vivo injection and electroporation of the embryonic rat neocortex provides a powerful tool for the manipulation of individual progenitors lining the walls of the lateral ventricle. This technique is now widely used to study the processes involved in corticogenesis by over-expressing or knocking down genes and observing the effects on cellular proliferation, migration, and differentiation. In comparison to traditional knockout strategies, in-utero electroporation provides a rapid means to manipulate a population of cells during a specific temporal window. In this video protocol, we outline the experimental methodology for preparing rats for surgery, exposing the uterine horns through laporatomy, injecting DNA into the lateral ventricles of the developing embryo, electroporating DNA into the progenitors lining the lateral wall, and caring for animals post-surgery. Our laboratory uses this protocol for surgeries on E15-E21 rats, however it is most commonly performed at E16 as shown in this video.

 

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Walantus W, Elias L, Kriegstein A (2007). In Utero Intraventricular Injection and Electroporation of E16 Rat Embryos. JoVE. 6. http://www.jove.com/index/details.stp?id=236, doi: 10.3791/236
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10/24/2007 2:50:58 PM
Barry responded with a statement of type: Neutral
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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10/29/2007 5:19:44 PM
dcurrle responded with a statement of type: Neutral
Try different methods of purifying the DNA.
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10/31/2007 2:12:06 AM
bioadventurist responded with a statement of type: Neutral
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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10/31/2007 8:53:59 AM
Barry responded with a statement of type: Neutral
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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05/28/2010 11:40:41 AM
m.barkats responded with a statement of type: Neutral
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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07/17/2008 2:59:20 PM
Derek responded with a statement of type: Neutral

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

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09/08/2009 1:23:03 AM
sara responded with a statement of type: Neutral
what are the medicine for motor neuron disease?
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11/10/2009 4:19:11 PM
Julie Cardin responded with a statement of type: Question
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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11/10/2009 4:40:55 PM
Anonymous responded with a statement of type: Neutral

BTX Harvard Apparatus Tweezertrodes, you can get them from Fisher and other vendors. -William
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08/07/2010 8:49:07 AM
blum61 responded with a statement of type: Neutral
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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08/07/2010 8:50:00 AM
blum61 responded with a statement of type: Question
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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07/19/2007

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doi: 10.3791/236 

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Journal of Visualized Experiments (JoVE) is an online research journal employing visualization to increase reproducibility and transparency in biological sciences.

 

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