FIG. 6.

FIG. 6.

Analysis of off-target effects of EGFP knockdown. (A) Principle component analysis of mRNA expression levels in all sequenced samples. The first two principle components are shown and are annotated by the percentage of the variance explained by each of those components. The colors differentiate the samples by experimental condition, while the shape by analysis time point. (B) Volcano plot showing the relation of the log2 transformed fold changes of read counts at 128-cell stage to their log10 transformed P-values adjusted for multiple comparison tests with the Benjamini and Hochberg correction, as implemented in DESeq2. The point originating from the EGFP transcript has been marked. The dashed red line shows the 0.1 adjusted p-value threshold. (C) Comparison of biological process gene ontology terms enrichment of the upregulated and downregulated genes. Dots represent the term enrichment with color coding: red indicates high enrichment and blue indicates low. The sizes of the dots represent the ratio of genes containing the relevant term within up or down-regulated groups to the total number of genes in that group. (D) The plots showing the log2 transformed fold change and its respective P-value from likelihood ratio test for the full model, containing both condition and batch factors, and the reduced model containing only the batch, fitted to the counts of the groups of genes sharing the same minimal number of complementary nucleotides to the crRNA. The error bars show the standard error of the estimate. With each required nucleotide less, new genes are added to the tested group. The bar plot in the uppermost panel depicts the size of the relevant group.