Dear Christopher,
Here are the settings we are using to electroporate Chlamy and which is
working well in our hands.
After treatment with autolysin, the cells are resuspended in TAP medium
containing saccharose (40 mM). We are using electroporation cuvettes
with a 4 mm gap. 250 µl of cells and 100 ng of DNA are incubated
directly in the cuvette for 10 min on ice. We use an exponential decay
pulse setted at 800V and 25µF.
The cells are then transfered for 18 hours in 10 mL of fresh TAP medium
under dim light before being applied to selective medium.
I hope it helps
Regards
David Dauvillée
UGSF-UMR 8576 CNRS
e 22/10/2015 19:03, chlamy-request from oat.bio.indiana.edu a écrit :
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>>> Today's Topics:
>> 1. Electroporation of Chlamydomonas reinhardtii
> (Christopher Applegate)
> 2. RE: Electroporation of Chlamydomonas reinhardtii (Original
> sent from incorrect e-mail) (Christopher Applegate)
>>> ----------------------------------------------------------------------
>> Message: 1
> Date: Wed, 21 Oct 2015 14:50:20 -0800
> From: Christopher Applegate <applegate.christopherr from gmail.com>
> Subject: [Chlamydomonas] Electroporation of Chlamydomonas reinhardtii
> To: chlamy from magpie.bio.indiana.edu> Message-ID:
> <CAM6NckcrnwxJ4kPQbxXSCMUS1QT3iKAZ73xVfZk-za1ksm4DKQ from mail.gmail.com>
> Content-Type: text/plain; charset=UTF-8
>> Good Day Ladies and Gentleman,
>> Has anyone had any success transforming Chlamy cells via electroporation
> using the Gene Pulser Xcell by Bio Rad? I am attempting to do so here at
> the University of Alaska Anchorage, but I have not had any luck finding any
> recommended setting for this machine. Thank you for your time and
> consideration. Have a great day.
>> Sincerely,
> Chris Applegate
> University of Alaska Anchorage
>>> ------------------------------
>> Message: 2
> Date: Wed, 21 Oct 2015 14:54:47 -0800
> From: Christopher Applegate <crapplegate from alaska.edu>
> Subject: [Chlamydomonas] RE: Electroporation of Chlamydomonas
> reinhardtii (Original sent from incorrect e-mail)
> To: chlamy from magpie.bio.indiana.edu> Message-ID:
> <CABhiJbjjAK9OKgtD7KZWcCo9xeoVXBM9-2UaBMV79Lj7bR+LZw from mail.gmail.com>
> Content-Type: text/plain; charset=UTF-8
>> Good Day Ladies and Gentleman,
>> Has anyone had any success transforming Chlamy cells via electroporation
> using the Gene Pulser Xcell by Bio Rad? I am attempting to do so here at
> the University of Alaska Anchorage, but I have not had any luck finding any
> recommended setting for this machine. Thank you for your time and
> consideration. Have a great day.
>> Sincerely,
> Chris Applegate
> University of Alaska Anchorage
>>> ------------------------------
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>Chlamy from net.bio.net>http://www.bio.net/biomail/listinfo/chlamy>> End of Chlamy Digest, Vol 85, Issue 1
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>
--
*David Dauvillée*
CR CNRS
Unité de Glycogiologie Structurale et Fonctionnelle
UMR 8576 CNRS-USTL
Bât C9
59650 Villeneuve d'Ascq
Phone : +33 3 20 43 48 73
Fax: +33 3 20 43 65 55
*David.Dauvillee from univ-lille1.fr*
