qRT-PCR of selected genes that were differentially expressed based on suppression subtractive hybridization.

<p>Relative quantification of mRNA expression (fold change) of (A) <i>pyruvate kinase</i> (<i>pk</i>, JZ347493), (B) <i>glutamine synthetase</i> (<i>gs</i>, JZ347462), (C) <i>fumarate hydratase</i> (<i>fh</i>, JZ347458), (D) <i>prolactin</i> (<i>prl</i>, JZ347487), (E) <i>phosphofructokinase</i> (<i>pfk</i>, JZ347479), (F) <i>Na⁺/K⁺-ATPase α2</i> (<i>nkaα2</i>, JZ347474), (G) <i>pyruvate kinase</i> (<i>pk</i>, JZ347493), (H) <i>prolactin</i> (<i>prl</i>, JZ347487) and (I) <i>ferritin heavy chain</i> (<i>fth</i>, JZ347374), using β-actin as the reference gene, in the brain of <i>Protopterus annectens</i> aestivated for 6 days (d) (A–F) or 6 months (G–I) in air with reference to the those of fish kept in freshwater as control. Results represent mean+S.E.M. (<i>N</i> = 6). *Significantly different from the corresponding freshwater control (<i>P</i><0.05).</p

Known transcripts found in the reverse library (down-regulation) obtained by suppression subtractive hybridization PCR from the brain of <i>Protopterus annectens</i> aestivated for 6 days in air with fish kept in freshwater as the reference for comparison.

<p>Known transcripts found in the reverse library (down-regulation) obtained by suppression subtractive hybridization PCR from the brain of <i>Protopterus annectens</i> aestivated for 6 days in air with fish kept in freshwater as the reference for comparison.</p

Primers used for quantitative real-time PCR on <i>acyl-CoA desaturase</i> (<i>acd</i>), <i>argininosuccinate synthetase 1</i> (<i>ass1</i>), <i>betaine-homocysteine S-methyltransferase 1</i> (<i>bhmt1</i>), <i>ceruloplasmin</i> (<i>cp</i>), <i>carbamoyl-phosphate synthetase III</i> (<i>cpsIII</i>), <i>fumarate hydratase</i> (<i>fh</i>), <i>ferritin light chain</i> (<i>ftl</i>), <i>glyceraldehyde-3-phosphate dehydrogenase</i> (<i>gapdh</i>), <i>superoxide dismutase 1</i> (<i>sod1</i>) from the liver of <i>Protopterus annectens</i>.

<p>Primers used for quantitative real-time PCR on <i>acyl-CoA desaturase</i> (<i>acd</i>), <i>argininosuccinate synthetase 1</i> (<i>ass1</i>), <i>betaine-homocysteine S-methyltransferase 1</i> (<i>bhmt1</i>), <i>ceruloplasmin</i> (<i>cp</i>), <i>carbamoyl-phosphate synthetase III</i> (<i>cpsIII</i>), <i>fumarate hydratase</i> (<i>fh</i>), <i>ferritin light chain</i> (<i>ftl</i>), <i>glyceraldehyde-3-phosphate dehydrogenase</i> (<i>gapdh</i>), <i>superoxide dismutase 1</i> (<i>sod1</i>) from the liver of <i>Protopterus annectens</i>.</p

Known transcripts found in the forward library (up-regulation) obtained by suppression subtractive hybridization PCR from the brain of <i>Protopterus annectens</i> aestivated for 6 months in air with fish kept in freshwater as the reference for comparison.

<p>Known transcripts found in the forward library (up-regulation) obtained by suppression subtractive hybridization PCR from the brain of <i>Protopterus annectens</i> aestivated for 6 months in air with fish kept in freshwater as the reference for comparison.</p

Primers used for quantitative real-time PCR on <i>fumarate hydratase</i> (<i>fh</i>), <i>ferritin heavy chain</i> (<i>fth</i>), <i>glutamine synthetase</i> (<i>gs</i>), <i>Na⁺/K⁺-ATPase α2</i> (<i>nkaα2</i>), <i>phosphofructokinase</i> (<i>pfk</i>), <i>pyruvate kinase</i> (<i>pk</i>) and <i>prolactin</i> (<i>prl</i>), with <i>β-actin</i> as the reference gene, from the brain of <i>Protopterus annectens</i>.

<p>Primers used for quantitative real-time PCR on <i>fumarate hydratase</i> (<i>fh</i>), <i>ferritin heavy chain</i> (<i>fth</i>), <i>glutamine synthetase</i> (<i>gs</i>), <i>Na⁺/K⁺-ATPase α2</i> (<i>nkaα2</i>), <i>phosphofructokinase</i> (<i>pfk</i>), <i>pyruvate kinase</i> (<i>pk</i>) and <i>prolactin</i> (<i>prl</i>), with <i>β-actin</i> as the reference gene, from the brain of <i>Protopterus annectens</i>.</p

Known transcripts found in the reverse SSH library (down-regulation) obtained by suppression subtractive hybridization PCR from the liver of <i>Protopterus annectens</i> after 1 day of arousal from 6 months of aestivation with fish aestivated for 6 months in air as the reference for comparison.

<p>Known transcripts found in the reverse SSH library (down-regulation) obtained by suppression subtractive hybridization PCR from the liver of <i>Protopterus annectens</i> after 1 day of arousal from 6 months of aestivation with fish aestivated for 6 months in air as the reference for comparison.</p

Known transcripts found in the forward library (up-regulation) obtained by suppression subtractive hybridization PCR from the liver of <i>Protopterus annectens</i> after 1 day of arousal from 6 months of aestivation with fish aestivated for 6 months in air as the reference for comparison.

<p>Known transcripts found in the forward library (up-regulation) obtained by suppression subtractive hybridization PCR from the liver of <i>Protopterus annectens</i> after 1 day of arousal from 6 months of aestivation with fish aestivated for 6 months in air as the reference for comparison.</p

Known transcripts found in the forward library (up-regulation) obtained by suppression subtractive hybridization PCR from the brain of <i>Protopterus annectens</i> aestivated for 6 days in air with fish kept in freshwater as the reference for comparison.

<p>Known transcripts found in the forward library (up-regulation) obtained by suppression subtractive hybridization PCR from the brain of <i>Protopterus annectens</i> aestivated for 6 days in air with fish kept in freshwater as the reference for comparison.</p

Known transcripts found in the forward library (up-regulation) obtained by suppression subtractive hybridization PCR from the liver of <i>Protopterus annectens</i> aestivated for 6 months in air with fish kept in fresh water as the reference for comparison.

<p>Known transcripts found in the forward library (up-regulation) obtained by suppression subtractive hybridization PCR from the liver of <i>Protopterus annectens</i> aestivated for 6 months in air with fish kept in fresh water as the reference for comparison.</p

Quantitative RT-PCR results of selected genes that were differentially expressed in the SSH libraries.

<p>Relative quantification of mRNA expression (fold change) of (A) <i>betaine-homocysteine S-methyltransferase 1</i> (<i>bhmt1</i>, JZ575536), (B) <i>fumarate hydratase</i> (<i>fh</i>, JZ575565), (C) <i>argininosuccinate synthetase 1</i> (<i>ass1</i>, JZ575533), (D) <i>carbamoyl-phosphate synthetase III</i> (<i>cpsIII</i>, JZ575539), (E) <i>superoxide dismutase 1</i> (<i>sod1</i>, JZ575606), (F) <i>ceruloplasmin</i> (<i>cp</i>, JZ575541), (G) <i>acyl-CoA desaturase</i> (<i>acd</i>, JZ575387), (H) <i>ferritin light chain</i> (<i>ftl</i>, JZ575418) and (I) <i>glyceraldehyde-3-phosphate dehydrogenase</i> (<i>gapdh</i>, JZ575429), using <i>β-actin</i> as the reference gene, in the liver of <i>Protopterus annectens</i> after 6 months (mon) of aestivation as compared with the freshwater control (A-F), or 1 day (d) of arousal from 6 mon aestivation as compared with fish aestivated for 6 mon (G-I). Results represent mean + S.E.M. (<i>N</i> = 6). *Significantly different from the corresponding control (<i>P</i><0.05).</p