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Once the universal standard gel was completed it was critical to determine if a
control gel would actually match the standard and to establish acceptable statistical limits
of matching. To test these aims a co-investigator (Archana Pandey) isolated and purified
protein from P. putida KT2440 grown to mid-log phase on 5mM succinic acid and
separated the sample through IEF-SDS-PAGE, as previously described. The resulting gel
was compared to the universal standard gel and evaluated based on the %CV and the
percent of spots that matched within the two gels. It was found that the %CV was 33%
and the percent of matching spots were 54% out of the total spots present on Padney’s
gel.
Previously reported studies found that gels created from identical samples led to
%CV values ranging from 19 to 26% and gels from different preparations of the same
sample resulted in 24 – 47% variability. [12] Based on these previous studies, it can be
assumed that future control gels can be quantitatively compared to the universal standard
gel. If the future control gels fall within this statistical range for %CV value (24 – 47%)
then it can be concluded that the gel agrees with the universal standard gel and that
differences in protein expression within the sibling gels are significant and not attributed
to technical variability.
Based on these findings the two gel sets that were previously discussed can now
be quantitatively compared to the universal gel standard. For this analysis only the
second version of the universal standard gel, the minimal requirements gel, will be the
basis of comparison. This analysis was based on three criteria; the first two being the
coefficient of variation and the percent of spots that could be matched between the gels.
The third variable is evaluating intensity changes within the matched protein spots (based
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