Table 1.
Genes and primer sequences of Calanus sinicus used in this study.
GAPDH, EF1ɑ and 16S are housekeeping genes, while the other six are target genes.
Fig 1.
Sampling station for Calanus sinicus.
The sampling station is marked with a star (☆) and is located inside the Yellow Sea Cold Water Mass (YSCWM). The boundary of the YSCWM is defined by the 10°C bottom isotherm and marked with bold black line.
Fig 2.
Vertical profiles of temperature and Chl a concentration at the Calanus sinicus sampling station.
Fig 3.
Oxygen consumption rate (OCR) of Calanus sinicus cultured under different temperatures by food environments.
9NF = 9°C not fed, 9F = 9°C fed, 18NF = 18°C not fed and 18F = 18°C fed.
Fig 4.
The composition time series of developmental stages and jaw phases in Calanus sinicus.
(a) 9NF (9°C not fed), (b) 18NF (18°C not fed), (c) 9F (9°C fed) and (d) 18F (18°C fed).
Fig 5.
Jaw phase morphology of Calanus sinicus.
(a) pre-apolysis on Day 0; (b) post-apolysis on Day 6 in Group 9F (9°C fed), tooth forming; (c) post-apolysis on Day 9 in Group 9F. Scale bars: 20 μm.
Fig 6.
Relative gene expression variation of genes associated with the molting process of Calanus sinicus.
(a) EcR (ecdysteroid receptor) expression. (b) FAMeT (Farnesoic acid O methyltransferase) expression. 9NF = 9°C not fed, 9F = 9°C fed, 18NF = 18°C not fed and 18F = 18°C fed. D0 to D16 represent Days 0 to 16. The expression level was normalized to the geometric mean of three housekeeping genes (GAPDH, 16s, EFα1). Error bars show the standard deviation. Significant differences (LSD): P < 0.05 (*); P < 0.01(**).
Fig 7.
Time series of gonad developmental stages in Calanus sinicus in different temperatures by food conditions.
(a) 9NF (9°C not fed), (b) 18NF (18°C not fed), (c) 9F (9°C fed) and (d) 18F (18°C fed). GS1 to GS4 represent gonad development stages 1–4.
Fig 8.
Gonad development states (GS2-GS4) in Calanus sinicus at the end of the culture experiment.
(a) Group 9NF (9°C not fed), GS2, developing oocytes (OS1 and OS2); (b) Group 9F (9°C fed), GS4, mature oocytes (OS4); (c) Group 18NF (18°C not fed); (d) Group 18F (18°C fed). Scale bar: 400 μm.
Fig 9.
Relative gene expression variation of gene Vitellogenin receptor (VgR) of Calanus sinicus.
9NF = 9°C not fed, 9F = 9°C fed, 18NF = 18°C not fed and 18F = 18°C fed. D0 to D16 represent Days 0 to 16. The expression level was normalized to the geometric mean of three housekeeping genes (GAPDH, 16s, EFα1). Error bars show the standard deviation. Significant differences (LSD): P < 0.05 (*); P < 0.01(**).
Fig 10.
Oil sac proportion (OSV%) of Calanus sinicus cultured in different temperatures by food conditions.
9NF = 9°C not fed, 9F = 9°C fed, 18NF = 18°C not fed and 18F = 18°C fed.
Fig 11.
Cumulative lipid costs for respiration and total lipid loss in oil sacs of Calanus sinicus.
(a) 9NF (9°C not fed), (b) 18NF (18°C not fed), (c) 9F (9°C fed) and (d) 18F (18°C fed).
Fig 12.
Relative gene expression variation of genes associated with lipid consumption of Calanus sinicus.
(a) HOAD (hydroxyacyl CoA dehydrogenase) expression. (b) DI (dienoyl-CoA isomerase) expression. 9NF = 9°C not fed, 9F = 9°C fed, 18NF = 18°C not fed and 18F = 18°C fed. D0 to D16 represent Days 0 to 16. The expression level was normalized to the geometric mean of three housekeeping genes (GAPDH, 16s, EFα1). Error bars show the standard deviations. Significant differences (LSD); P < 0.05 (*), P < 0.01(**).
Fig 13.
Relative gene expression variation of gene ferritin of Calanus sinicus.
9NF = 9°C not fed, 9F = 9°C fed, 18NF = 18°C not fed and 18F = 18°C fed. D0 to D16 represent Days 0 to 16. The expression level was normalized to the geometric mean of three housekeeping genes (GAPDH, 16s, EFα1). Error bars show the standard deviations. Significant differences: LSD, p <0.05 (*), p <0.01(**).
Fig 14.
Principal component analysis of the relative gene expression data of Calanus sinicus in Group 18F (18°C fed).
(a) The loading plot of six genes. (b) Scatterplot of the averages of the first two principal component scores for C. sinicus. D0 to 16 represent Days 0 to 16.
Fig 15.
Termination and subsequent development of the over-summering Calanus sinicus cultured in Group 18F (18°C fed).
The internal processes were divided into four periods: quiescence (Day 0), termination and molting process (Days 1–6), early gonad development (Days 6–9) and final gonad development (Days 9–16). The associated metabolic rates (OCR, dotted line), lipid reserve (OSV%, solid line) and gene expression patterns are shown. See Table 1 for gene abbreviations.