The samples for RNA analysis were harvested from the fermentors during the mid-log and late-log phase. The time points and dry cell weight of the mid-log and late-log phase can be seen in (Additional file 1: Table S1).
RNA-seq analysis An analysis of variance (ANOVA) was conducted on each of the independent variables separately: strain, Populus hydrolysate concentration, and time. Differentially expressed genes were defined as a selleck inhibitor 2-fold change in expression with a false discovery rate of less than 5% (p < 0.05). Of the 3,236 genes Selleckchem Ipatasertib in C. thermocellum, roughly 18% (n = 574) showed a difference in expression between strains. Furthermore, approximately 16% (n = 505) of the genes showed a change in expression between the three concentration comparisons. None of the genes showed a change in expression between the two time Quizartinib points. Since, there were
no statistically significant changes in expression of individual genes between the mid-log and late-log time points, the analysis considered-between strain and between-hydrolysate-concentration comparisons to be significantly different if the expression differences were significant for either of these two time points. Simple comparisons only consider the differences in gene expression from changing one of the three variables at a time: strain, Populus hydrolysate concentration or time. The ANOVA of the three independent variables in combination revealed approximately
55% (n = 1795) of the genes were differentially expressed in at least one of the simple comparisons (Additional file 2). Two types of analyses are the focus of this paper. The first analysis compares gene expression in the WT and PM strains in 0% v/v and 10% v/v Populus hydrolysate. A positive differential expression (upregulation) represents a higher expression level in the PM strain and a negative differential expression (downregulation) represents a lower expression level in the PM strain when compared to the WT strain. The second type of analysis compares gene expression under different concentrations of Populus hydrolysate within a given strain as follows: the PM in 0% versus 10% v/v Populus hydrolysate and 0% versus 17.5% v/v Populus hydrolysate, RVX-208 and the WT in 0% versus 10% v/v Populus hydrolysate. For these comparisons a positive differential expression (upregulation) represents an increase in expression level and a negative differential expression (downregulation) represents a decrease in expression level in the Populus hydrolysate compared to standard medium. Of the 1795 differentially expressed genes, 1740 are represented by these four comparisons. The remaining 55 genes are differentially expressed between the comparisons of the PM in 10% versus 17.5% v/v Populus hydrolysate or between the mid-log versus late-late log time points for a given condition.