For each dilution step a primary dispersion of 5 mg oil per liter was continuously diluted with seawater using 3 way solenoid valves. Nordtug et al. describe the procedures employed for generation Inhibitors,Modulators,Libraries of Inhibitors,Modulators,Libraries oil dispersions and dilutions, and the layout of the exposure chamber system. The ex posure containers consisted of 5 L borosilicate glass bot tles with bottoms removed and placed upside down in a water bath. Exposure solution and clean seawater was added to the lower part of the ex posure container through Teflon tubing. Water was drained from the surface through a 300 um plankton mesh. The temperature was con trolled by submerging all exposure chambers into a water bath. The flow through in all exposure units was kept con stant at 17,3 mL SD 1. 3 mL min, corresponding to mean residence time of the water of approximately 4.

5 hours. Dispersions were added by passive flow through thin inlet Teflon tubes and the flow was adjusted by changing the height of the inlet water column. A total number of 300 cod larvae were carefully intro duced into each exposure chamber. The larvae were fed live rotifiers which were added in batches Carfilzomib three times a day together with algae. Four parallel exposure chambers were used in order to achieve biological replicates for every exposure concentra tion. The exposure design used in the experiment is given in Figure 1A. The cod larvae were exposed for 96 hours, counted and sampled. Inhibitors,Modulators,Libraries Larvae sampling and RNA extraction At the end of the exposure period, the cod lar vae were immediately rinsed with distilled water and blot ting paper and flash frozen in liquid nitrogen, and stored at ?80 C before RNA isolation.

To ensure enough RNA was available for downstream transcriptomic analysis, 15 larvae were pooled together from each exposure unit. With this design, we had four biological replicates for each treatment, consisting of a total of 60 larvae. Due to an extensive sampling program and high lethality the available live larvae from one of the exposure units of the MDH and MDM groups Inhibitors,Modulators,Libraries was too few to be analyzed, leav ing only 3 parallels for these groups. In total, RNA for transcriptomic analyses was isolated from 26 samples. The larvae were thoroughly homogenized before RNA extraction using zirconium beads in a Precellys 24 homogenizer by ceramic beads CK28. Total RNA from Atlantic cod larvae was extracted using phenol chloroform extraction and Qiazol with a modified isopropanol precipitation step previously described elsewhere. The samples were sub sequently treated with DNA free, according to the manufacturers instructions and eluted in 50 uL RNase free MilliQ H2O. The RNA was then stored at ?80 C be fore further processing.