Biology Reference
In-Depth Information
and 100
L of 10 % (w/v) SDS. Immediately prior to pouring
the gel into the hand cast for Mini-PROTEAN two-gel elec-
trophoresis system (BioRad), add 50
μ
μ
L of 10 % (w/v) APS
L of TEMED and mix gently. The use of electrophore-
sis to remove it would lead to a reduced loss of these proteins.
10. More details about 1-DE and 2-DE separation methods are
described in two excellent reviews [
23
,
48
].
11. The condition of protein focusing must be optimized for each
study method. In our case, we use the PROTEAN IEF cell by
Bio-Rad. The conditioning phase involves the application of
previous steps at low voltage that allow to remove ions and
other contaminants containing the sample, and that interfere
on protein focusing. The current should not exceed 50
and 5
μ
A per
strips. For more information see the 2-D Electrophoresis for
Proteomics Manual by Bio-Rad.
12. More details about the MS analysis are described in [
56
,
64
,
71
-
73
].
13. The Botrytis cinerea Database was downloaded from the
Botrytis cinerea
Sequencing Project, Broad Institute of
Harvard and MIT (
http://www.broadinstitute.org/annota-
details about gel-free analysis are described in [
59
,
74
,
75
].
μ
Acknowledgements
This work was supported by the Spanish Ministry of Science and
Innovation (BotBank Project, EUI2008-03686), the Andalusian
Regional Government (Junta de Andalucía), and the University of
Córdoba (AGR-0164: Agricultural and Plant Biochemistry and
Proteomics Research Group).
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