mohammad tabish ahmed - eTheses Repository - University of ...

Chapter 3
of MGM100. The use of Tab21 did not have any affects on the overall complementation data
and similar levels of protein expression were obtained. It was concluded that, as was the case
with MGM100, the Cpn60.2 chaperonin complements for the loss of endogenous GroEL in
Tab21, while Cpn60.1 does not. A summary of the complementation and expression results
can be seen in table 3.2 and figure 3.6.
As discussed above, since Tab21 is a protein expression strain that makes use of the T7 RNA
polymerase expression system, the next step in trying to achieve higher levels of protein
expression of Cpn60.1 involved the transfer of the groES-cpn60.1 operon and the cpn10-
cpn60.1 operon onto a pET plasmid (section 3.3.1).
1 2 3
72 kDa
55 kDa
Fig 3.6: SDS-PAGE gel showing over-expression of Cpn60.1 and Cpn60.2 in Tab21. All
the samples were taken from cultures after 5 hours incubation with 0.1 mM IPTG and 0.2%
arabinose.
Lane 1- Prestained protein marker (Fermentas)
Lane 3- Tab21/ pTrc-GroES-Cpn60.2
Lane 2- Tab21/ pTrc-GroES-Cpn60.1
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