# Abscisic Acid--Defensive Player in Flax Response to _Fusarium culmorum_ Infection
## 3 Results
### Analysis of Selected Isoprenoid Levels  
We measured the amounts of selected isoprenoids in the flax tissue treated with _F. culmorum_ at 6 hpi, 12 hpi, 24 hpi, 36 hpi, and 48 hpi. The total level of phytosterols was somewhat (ca. 5%) elevated in the measured time-points (except the 6 hpi, where it was 78.4% of the control). Particular phytosterols were quantified and are presented in Figure 4A. In general, decreased levels of all measured phytosterols were observed at 6 hpi and slight, but consistent increase at 36 hpi and 48 hpi.  
Similar as it was in the case of sterols, the levels of all measured carotenoids, as well as chlorophylls were down to 85% of the control on average at the beginning of the infection, but then their levels increased to reach maximum 1.24-fold of the control for violaxanthin at 12 hpi, 1.11-fold of the control for tocopheroids at 24 hpi to drop down to the control level at 36 hpi. Then, at 48 hpi, a decrease was noted for all of the compounds in this group, by 12% compared with the control on average (Figure 4B). The level of ABA was considerably increased throughout all the time-points investigated and reached maximally 7.6-fold of the control at 12 hpi (Figure 4C).  
Figure 4: Heatmap of terpenoid metabolites in flax after _F. culmorum_ infection (x–fold of the control). (**A**)— phytosterols; (**B**)—carotenoids; (**C**)—ABA. Detailed results on the expression can be found in Supplementary File S4.  
Analysis of the Expression Level of Callose Synthase Gene and Callose Content in Response to F. culmorum Infection  
ABA is considered a regulator in plants' response to infection and influences many of defense genes that are involved in the biosynthesis of compounds participating in the resistance to pathogens, such as callose. We measured the transcript levels of callose synthase genes (Figure 5). Although inconsistent, the transcription of \(CALS1\) (callose synthase 1) gene was elevated after the infection reaching its maximum at 48 hpi (2.03-fold of the control). Similarly, the level of \(CALS3\) (callose synthase 3) gene was increased in the measured time-points and the increase was most prominent at 36 hpi and 48 hpi (2.17- and 2.07-fold of the control, respectively). \(CALS2\) (callose synthase 2) gene transcript level was virtually at the control level in all time-points with somewhat increase at 48 hpi (1.39-fold of the control). \(CALS4\) (callose synthase 4) gene expression decreased at 24 hpi and 36 hpi (0.68- and 0.31-fold of the control, respectively) to return to the control level at 48 hpi.  
We measured the amounts of callose at 6 hpi, 12 hpi, 24 hpi, 36 hpi, and 48 hpi. After a decrease observed in 24 hpi to 0.69-fold of the control, its level started to grow up to reach 2.15-fold of the control at 48 hpi (Figure 6).  
Figure 5: Transcript levels of callose synthase genes (\(CALS1\), \(CALS2\), \(CALS3\), \(CALS4\)) in flax seedlings treated with _F. culmorum_ measured at five timepoints after the infection. Results are presented as a fold of the control (means of three biological repeats \(\pm\) sd). Statistically significant changes are marked with asterisks.