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Western blot assays confirmed the absence of Ga5DH in the deletion mutant and the return of Ga5DH expression in the complemented strain (Fig. 1B).
The peroxidase activity was restored in the complemented strain.
The reduced tolerance to H2O2 and NO of the mutant was recovered in the complemented strain.
The parental level of virulence was significantly restored in the complemented strain (p<0.001).
As expected, the cytotoxicity level was fully restored in the complemented strain MF213 (p-value>0.05) in the three hosts.
The parental levels of the CFU counts from lungs and spleens were recovered in the complemented strain.
Similar(31)
Western analyses were performed to confirm the expression of the proteins in the complemented strains.
In the complemented strains, the proteins SpiC and SpvB are expressed from the pXG0 low copy plasmid, under the control of the respective promoter.
The wild type level of association was restored in the complemented strains confirming the effect was due to the absence of the LPS polysaccharides rather than a secondary mutation.
Next we compared the expression level of all three genes in the complemented strains using RT-qPCR.
Neither Myc11 pMs100 nor Myc11 pMs101 produced this unique lipid, and the native GPL distribution was almost completely restored in the complemented strains, although the relative amounts were not identical to that of the wild type.
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Justyna Jupowicz-Kozak
CEO of Professional Science Editing for Scientists @ prosciediting.com