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To confirm that Vhl alleles were indeed deleted in Glu-Cre;Vhl f/f mice, we isolated α-cells, β-cells and endothelial cells from pancreatic islets via flow cytometry, and obtained genomic DNA for each cell population.
To confirm that the Vhl alleles were indeed deleted in the pancreas of Pdx1-Cre;Vhl f/f newborn pups, whole pup body DNA and pancreatic DNA were isolated by laser capture microdissection.
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Lastly, qPCR was performed using genomic DNA from sorted splenic CD4+Foxp3-GFP+ cells to assess the efficiency of ppp4c deletion in the CD4cre:PP4f/f:Foxp3-GFP+ mice; the results showed that the ppp4c gene was indeed deleted in >85% of the Treg cells.
We confirmed that the RanBP2/RanGAP1 complex is indeed depleted from kinetochores when Ska3/Rama1 is deleted in HeLa cells.
The strain ∆rha1∆lra3 did not show any growth on l-rhamnose (data not shown) nor l-rhamnose consumption under these conditions (Fig. 2) indicating that the genes deleted were indeed required for l-rhamnose catabolism in A. niger.
It is of note that conversely Myc-N amplification and 1p deletion were indeed correlated, being 72.7% of Myc-N amplified patients also deleted of 1P.
The psychometric properties of the OLQ-11 (obtained by deleting the two items) were indeed better, in particular the exploratory factor solution.
These results showed that the four deleted genes in the mutants were indeed responsible for their inability to grow as typical "fried egg" L-form colonies.
The observed phenotype is indeed caused by deleting rbbA or yhjD because a pBAD-plasmid expressing RbbA or YhjD under the control of an arabinose-inducible promoter, but not a control plasmid without the respective genes, restored the antibiotic resistance of rbbA or yhjD null strains to wild type levels (Figure 4F; only rbbA complementation is shown).
To determine whether the genes that are heterozygously deleted are indeed dosage-sensitive at the transcript level, we focused on genes expressed in brain and carried out real-time quantitative RT-PCR analyses on brain tissue.
The presented results do however provide further evidence that human and mouse sperm potassium currents are indeed different, and should not be deleted.
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