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Corrections for multiple sampling (Bonferroni method and Dunnett's multiple comparison test) were used where appropriate.
Since it is not statistically valid to test multiple divisions with corrections for multiple sampling, rigorous statistical evaluation is achieved by defining divisions in a training set and then validating them in an independent non-overlapping validation set.
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Comparisons between means were performed using one-tailed Student's t-test for paired data with Bonferroni correction for multiple sampling as appropriate.
This involved linear modeling for each gene, use of an empirical Bayes method to moderate the standard errors of the estimated log-fold changes, and correction for multiple sampling using the method of Benjamani and Hochberg [ 34].
Pair-wise comparison analyses of near-triploid (n = 22) versus near-diploid/tetraploid (n = 27) NBTs revealed small sets of differentially expressed genes when using a stringent correction for multiple sampling, (6 genes [FDR < 0.01] and 12 genes [SDP < 0.1]) (see Additional file 2).
These were: (1) genes with a more than twofold difference in mean expression between control and exposed samples; (2) genes with significantly different (p < 0.05) expression between control and exposed samples after correction for multiple sample testing [ 56].
Statistical analysis was performed with Stata 9.1 for Windows (StataCorp LP, College Station, TX, USA), by using one-way analysis-of-variance model with Bonferroni multiple-comparison correction for multiple sample experiments and the Mann–Whitney test for experiments with comparison between two groups.
The fact that the DAVID pathway enrichment analysis system itself includes an algorithm for multiple sample correction effectively limits type 1 errors during identification of enriched pathways.
Further, the potential for type I error in identifying overrepresented pathways is also controlled for by the fact that the Database for Annotation, Visualization and Integrated Discovery (DAVID) tool that was used for enrichment analysis itself includes an algorithm for multiple sample correction.
In tests for Hardy Weinberg equilibrium (HWE) at individual loci (Supplementary Information, Table S3), two tests out of 832 were significant after corrections for multiple tests and all samples except 08Salmon_W were in HWE in tests over all loci (Table 1).
The results presented here are from a single sample and do not withstand conservative corrections for multiple comparisons.
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