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This so-called high transformation background is supposedly caused by cell stress and cell rupture.
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This transformation included background correction, normalization and summarization of expression values using Brainarray version 15 custom chip definition (cdf) files generated with the Ensembl annotation set [ 17].
For datasets using Ambion miRNA chips and OSU-CCC chips, we used vsn transformation after background subtraction [ 39] and applied quantile normalization method [ 40].
After log2-transformation and background subtraction, data were normalized to a median intensity ratio of one.
In addition, to be able to take the log base 2 transformation of the background adjusted intensity values, all background adjusted intensity values below zero were replaced with the value 0.00001.
In contrast, the 208F cells showed no background transformation and allowed us to detect a low frequency of transformation by RON and Stk.
This results in a smooth monotonic transformation of the background subtracted intensities such that all the corrected intensities are positive.
We used log2 transformation and without background correction.
We perform this transformation for each background window to reduce the effect of outlier positions associated with a large number of tags.
Unless a very flat subclone of NIH 3T3 cells is used in these assays, as we used here, a high and variable background transformation rate can be observed.
These reports showed a very high background transformation rate in the NIH 3T3 cells (50 foci per μg control DNA, ref. [ 24]; see photographs of cell culture dishes in Fig. 2 of ref. [ 23]) casting doubt on the reliability of the transformation assay.
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