Summary - Survey of the "ideal" protein-TAG

HAUSER MARIE-THERES hauser at edv2.boku.ac.at
Thu Feb 21 15:04:07 EST 2002


Dear collegues,

Thanks a lot for the answers. If there are more comments please
forward them to me - I will post them again.

Todate following answers about different tag experiences can be
summarizes as follows:

Concering Western blotting:

very clean results with 5xc-Myc or single HA-tagged proteins in
Arabidopsis. A triple HA-tag will give you even better signal. I have
had no succcess with the 6xHIS tag; I cannnot detect the same
protein I see clearly with the HA tag. anti-myc Abs (9E10) come
from Santa Cruz Biotechnology; ant-HA (high affinity, 3F10) from
Roche.

Good results with single HA-tag. Imporant was the position of the
HA-tag. In the middle of the protein - nothing to see on the Western.
On the C-terminus - good Western results.
Anti-HA and anti-myc antibodies from Roche (former Boehringer
Mannheim) are good.
2.5x - 6x myc-tag work well but not 1x myc-tags.

we use HA and c-myc successfully for westerns. HA is detected
with a high affinity monoclonal from the Company-formally-known
as-Boehringer; c-myc is detected with a polyclonal from Santa
Cruz.
I have heard bad reports with 6xHis antibodies but don't know
details.

Concerning IP:
  the anti-HA (3F10) beads from Roche work very well in IP.


The original question was:
I am wondering if somebody has experience with different TAGs in
Arabidopsis for
a.) Western Blotting
b.) Immunoprecipitation
c.) Immunocytchemistry
and which are the best antibodies. Which company sells them?
Who used successfully the HA, c-myc, FLAG, His, T7, AU1, AU5,
KT3, Glu-Glu, VSV-G, V5, ........?
Thanks a lot for your help.
Yours,
Marie-Theres




----------------

Dr. Marie-Theres Hauser

Center of Applied Genetics
University of Agricultural Sciences Vienna
Muthgasse 18
A-1190 Vienna
Austria/EUROPE

Tel: +43-1-36006-6371
FAX: +43-1-36006-6392
e-mail: hauser at mail.boku.ac.at
http://www.boku.ac.at/zag/AG_hauser.htm

----------------

Dr. Marie-Theres Hauser

Center of Applied Genetics
University of Agricultural Sciences Vienna
Muthgasse 18
A-1190 Vienna
Austria/EUROPE

Tel: +43-1-36006-6371
FAX: +43-1-36006-6392
e-mail: hauser at mail.boku.ac.at
http://www.boku.ac.at/zag/AG_hauser.htm

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