Table 2. Classification ability of the normoxia and hypoxia groups for 24 samples used in RNA-seq, and across the smoltification process of Chinook salmon, and the distressed (moribund/recently dead) fish using the hypoxia biomarkers. Biomarkers identified by Shrunken Centroid method for each group were placed into a linear discriminant analysis (LDA) for classifying groups within each treatment and mortality. LDA training set used two-thirds of the entire dataset. Presented are the classifications on the remaining one-third testing set. Columns are the LDA classification groups and rows are the real group. Salinity symbols are FW for freshwater, BW for brackish, and SW for seawater groups. Dissolved oxygen symbols are N for normoxia and H for hypoxia groups. Fish size (mean ± SD) symbols are L for length and W for weight.

Treatment	Sample size		Fish size				LDA Classification ability (%)			Significant genes identified by Shrunken analysis	Function of genes
	Training	Testing	L (cm)		W (g)		N	H	Average
			N	H	N	H
RNA-seq samples	16	8	8.4 ± 1.2	8.7 ± 1.4	6.5 ± 3.0	7.1 ± 3.4	100.0	75.0	87.5	Claspin, Tonsl, ARHGAP11A, KIF2C, AURKB, ERCC6L	DNA synthesis, cell division, and cell cycle
										RAMP1	Ion transport
Pre-smolt
FW	14	7	5.2 ± 0.4	5.2 ± 0.3	1.5 ± 0.3	1.5 ± 0.2	66.7	75.0	70.8	Ndc80, Kif4, Anillin, Ncapd3	DNA synthesis, cell division, and cell cycle
BW	12	6	5.2 ± 0.3	5.2 ± 0.3	1.5 ± 0.2	1.4 ± 0.2	66.7	66.7	66.7	Tonsl, NCAPG2, Anillin, KIF2C, CIT, Claspin, PCNA, Rab9, Lmnb1	DNA synthesis, cell division, and cell cycle
										SOX.5	Cell cycle arrest
										ACOT9	Oxidative stress
SW	13	6	5.2 ± 0.3	5.1 ± 0.3	1.7 ± 0.3	1.4 ± 0.2	75.0	100.0	87.5	Kif15, Anillin	DNA synthesis, cell division, and cell cycle
										CDKN1B	Cell cycle arrest
										RAMP1	Ion transport
										GPX3	Oxidative stress
										PAM	Peptide amidation
Smolt
FW	14	8	7.6 ± 0.4	7.2 ± 0.3	4.6 ± 0.8	4.1 ± 0.6	75.0	75.0	75.0	PCNA, Rab9, Kif4, Claspin, Ncapd3, Ndc80	DNA synthesis, cell division, and cell cycle
										SOX.5	Cell cycle arrest
										GPX3	Oxidative s tress
										ECE.2	Neuropeptide processing
BW	12	6	7.5 ± 0.5	7.5 ± 0.4	4.6 ± 1.0	4.6 ± 0.8	100.0	33.0	66.7	Lmnb1, FAM214A	DNA synthesis, cell division, and cell cycle
										CDKN1B	Cell cycle arrest
										RAMP1	Ion transport
SW	12	6	7.6 ± 0.2	7.5 ± 0.5	4.7 ± 0.6	4.6 ± 1.0	66.7	100.0	83.3	Anillin, H2AFV, RRM1	DNA synthesis, cell division, and cell cycle
										CDKN1B	Cell cycle arrest
										RAMP1	Ion transport
De-smolt
FW	12	6	9.5 ± 0.9	9.0 ± 1.3	9.0 ± 2.7	7.7 ± 2.8	66.7	100.0	83.3	Ndc80, Kif4, Anillin, ARHGAP11A, KIF2C, AURKB	DNA synthesis, cell division, and cell cycle
BW	20	10	9.5 ± 1.1	8.9 ± 1.0	7.8 ± 2.7	7.5 ± 2.6	40.0	60.0	50.0	FAM214A	DNA synthesis, cell division, and cell cycle
										ACOT9	Oxidative stress
										RAMP1	Ion transport
SW	31	16	7.6 ± 1.2	8.5 ± 1.0	6.8 ± 2.8	6.6 ± 2.4	100.0	75.0	87.5	Claspin, RRM1, Tonsl, FAM214A, Anillin, Rab9, CIT, Lmnb1	DNA synthesis, cell division, and cell cycle
										CDKN1B, SOX.5, MFHAS1	Cell cycle arrest
										Calphotin	Cellular calcium regulation
										GPX3, ACOT9	Oxidative stress
										RAMP1	Ion transport
Distressed
FW	17	9	8.3 ± 0.9	8.0 ± 1.0	6.1 ± 2.1	5.5 ± 2.2	80.0	100.0	90.0	KIF2C	Cell division and DNA synthesis
										MFHAS1	Cell cycle arrest
										GPX3	Oxidative stress
										ECE.2	Neuropeptide processing
SW	34	17	7.3 ± 1.8	6.7 ± 1.7	4.4 ± 2.9	3.4 ± 2.3	77.8	100.0	88.9	FAM214A, Kif4, ARHGAP11A, H2AFV, Claspin, Kif15	Cell division and DNA synthesis
										SOX.5, MFHAS1, CDKN1B	Cell cycle arrest
										GPX3, ACOT9	Oxidative stress
										ECE.2	Neuropeptide processing
										RAMP1	Ion transport
										PAM	Peptide amidation