IC4R006-Microarray-2014-24913626 - Revision history

Xysj1988: /* Research Findings */

2016-06-22T13:03:33Z

‎Research Findings

Xysj1988: /* Research Findings */

2016-06-22T13:03:24Z

‎Research Findings

Xysj1988: /* Plant Materials & Treatment */

2016-06-22T13:02:41Z

‎Plant Materials & Treatment

Xysj1988: /* Plant Materials & Treatment */

2016-06-22T13:02:32Z

‎Plant Materials & Treatment

Xysj1988: /* The Background of This Project */

2016-06-22T13:02:25Z

‎The Background of This Project

Xysj1988: /* The Background of This Project */

2016-06-22T13:02:17Z

‎The Background of This Project

Xysj1988: /* The Background of This Project */

2016-06-22T13:02:04Z

‎The Background of This Project

Xysj1988: /* The Background of This Project */

2016-06-22T13:01:51Z

‎The Background of This Project

Xysj1988: /* The Background of This Project */

2016-06-22T13:01:29Z

‎The Background of This Project

Xysj1988: /* The Background of This Project */

2016-06-22T13:00:20Z

‎The Background of This Project

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 ==Research Findings==
-[[File: IC4R006-Microarray-2014-24913626-3.png|center|thumb|527px|'''Figure 4. Expression of selected suberin biosynthesis genes in the outer part of root (OPR), cortical parenchyma (CP), and central cylinder (CC) of roots of rice.''']]
+[[File: IC4R006-Microarray-2014-24913626-3.png|right|thumb|527px|'''Figure 4. Expression of selected suberin biosynthesis genes in the outer part of root (OPR), cortical parenchyma (CP), and central cylinder (CC) of roots of rice.''']]
 * As a result, the signal intensities of 167 (0.4%) of the 41 880 gene probes spotted on the microarray slide were significantly higher or lower in OPR tissue forming the barrier to ROL than in OPR tissues that were not forming a barrier. Of the 167 gene probes, 125 (0.3% of the probes on the array) were upregulated and 42 (0.1% of the probes on the array) were downregulated (Fig. 2). After excluding homolo- gous gene probes, 98 genes were upregulated  and 30 genes were downregulated  under conditions inducing ROL barrier formation in the OPR of the basal part of long adventitious roots in stagnant deoxygenated nutrient solution. The most promi- nent GO classifications of the up- and downregulated genes were metabolic and cellular processes.
 * To confirm the key microarray results, this work selected 13 genes (11 upregulated genes that were predicted to be involved in suberin biosynthesis and two downregulated genes) for semiquantitative RT-PCR using RNA obtained from the laser microdissection-iso- lated OPR from long adventitious roots grown in stagnant deoxygenated condition. The 11 upregulated genes were predicted to be involved in suberin biosynthesis. One of these 11 genes, a POD gene (LOC_Os06g16350) is involved in the biosynthesis of both suberin and lignin. The two downregulated genes were PHOSPHOENOLPYRUVATE CARBOXYLASE (PEPC) (Lin et  al., 2004), which sup- plies oxaloacetate to the TCA cycle and CHAPERONIN 60/TCP-1 FAMILY PROTEIN (CPN60), which is required for correct protein folding. The expression of each of these genes in the OPR in long adventitious roots under stagnant deoxygenated conditions were upregulated or downregulated in agreement with the microarray findings.

@@ Line 17: / Line 17: @@
 ==Research Findings==
 * As a result, the signal intensities of 167 (0.4%) of the 41 880 gene probes spotted on the microarray slide were significantly higher or lower in OPR tissue forming the barrier to ROL than in OPR tissues that were not forming a barrier. Of the 167 gene probes, 125 (0.3% of the probes on the array) were upregulated and 42 (0.1% of the probes on the array) were downregulated (Fig. 2). After excluding homolo- gous gene probes, 98 genes were upregulated  and 30 genes were downregulated  under conditions inducing ROL barrier formation in the OPR of the basal part of long adventitious roots in stagnant deoxygenated nutrient solution. The most promi- nent GO classifications of the up- and downregulated genes were metabolic and cellular processes.
 * To confirm the key microarray results, this work selected 13 genes (11 upregulated genes that were predicted to be involved in suberin biosynthesis and two downregulated genes) for semiquantitative RT-PCR using RNA obtained from the laser microdissection-iso- lated OPR from long adventitious roots grown in stagnant deoxygenated condition. The 11 upregulated genes were predicted to be involved in suberin biosynthesis. One of these 11 genes, a POD gene (LOC_Os06g16350) is involved in the biosynthesis of both suberin and lignin. The two downregulated genes were PHOSPHOENOLPYRUVATE CARBOXYLASE (PEPC) (Lin et  al., 2004), which sup- plies oxaloacetate to the TCA cycle and CHAPERONIN 60/TCP-1 FAMILY PROTEIN (CPN60), which is required for correct protein folding. The expression of each of these genes in the OPR in long adventitious roots under stagnant deoxygenated conditions were upregulated or downregulated in agreement with the microarray findings.

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 * In each experiment, pots were arranged in a completely randomized design. 23-d-old plants were either continued in aerated solution or transplanted into N 2 -flushed or stagnant deoxygenated solution for 9 h (Fig. 1C). In N 2 -flushed nutrient solution, oxygen level was kept hypoxic (dissolved oxygen <1.0 mg l –1 ). Stagnant solution contained 0.1% (w/v) dissolved agar and was deoxygenated (dissolved oxygen <1.0 mg l –1 ) prior to use by preflushing with N 2 gas. The dilute agar prevents convective movements in solution (‘stagnant’ treatment) so this treatment mimics better than other solution culture methods the changes in gas composition found in waterlogged soils (e.g. decreased oxygen, increased ethylene) (Wiengweera et al., 1997).
 * Adventitious roots of 23-d-old plants were classified as short (65–85 mm) or long (115–135 mm), based on the length of the main axis at commencement of treatments. Selected short and long adventitious roots were marked near the base using small loops of sewing cotton, so these could be recognized over time.
-[[File: IC4R006-Microarray-2014-24913626-2.png|center|thumb|727px|'''Figure 2. Numbers of up- and downregulated gene probes at the outer part of root (OPR) during formation of a radial oxygen loss (ROL) barrier in rice (i.e. the OPR along the basal part of long adventitious roots, 9 h treatment in deoxygenated stagnant 0.1% agar nutrient solution).''']]
+[[File: IC4R006-Microarray-2014-24913626-2.png|center|thumb|927px|'''Figure 2. Numbers of up- and downregulated gene probes at the outer part of root (OPR) during formation of a radial oxygen loss (ROL) barrier in rice (i.e. the OPR along the basal part of long adventitious roots, 9 h treatment in deoxygenated stagnant 0.1% agar nutrient solution).''']]
 ==Research Findings==

@@ Line 14: / Line 14: @@
 * In each experiment, pots were arranged in a completely randomized design. 23-d-old plants were either continued in aerated solution or transplanted into N 2 -flushed or stagnant deoxygenated solution for 9 h (Fig. 1C). In N 2 -flushed nutrient solution, oxygen level was kept hypoxic (dissolved oxygen <1.0 mg l –1 ). Stagnant solution contained 0.1% (w/v) dissolved agar and was deoxygenated (dissolved oxygen <1.0 mg l –1 ) prior to use by preflushing with N 2 gas. The dilute agar prevents convective movements in solution (‘stagnant’ treatment) so this treatment mimics better than other solution culture methods the changes in gas composition found in waterlogged soils (e.g. decreased oxygen, increased ethylene) (Wiengweera et al., 1997).
 * Adventitious roots of 23-d-old plants were classified as short (65–85 mm) or long (115–135 mm), based on the length of the main axis at commencement of treatments. Selected short and long adventitious roots were marked near the base using small loops of sewing cotton, so these could be recognized over time.
 ==Research Findings==

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 * Another key feature contributing to waterlogging tolerance is aerenchyma formation. In rice roots, aerenchyma is formed constitutively, but the amount can be enhanced by soil water-logging (Armstrong, 1971; Pradhan et al., 1973; Jackson and Armstrong, 1999), low oxygen (Colmer et al., 2006), and ethylene (Justin and Armstrong, 1991; Colmer et al., 2006). On the other hand, the barrier to ROL is inducible in rice roots, forming in stagnant or waterlogged conditions, but not (or only weakly) in well-drained or aerated conditions (Colmer et al., 1998; Colmer, 2003a; Colmer et al., 2006; Insalud et al., 2006; Kotula et al., 2009; Shiono et al., 2011). Interestingly, low oxygen, ethylene, and elevated CO 2 , which arise from natural waterlogged soil, are not involved in triggering for- mation of the barrier to ROL in roots of rice (Colmer et al., 2006).
 * The signal that triggers formation of the barrier to ROL is not known yet, but it may involve root exudates, cellular degradation products (Armstrong and Armstrong, 2001; Voesenek and Sasidharan, 2013) or phytotoxins present in chemically reduced waterlogged soils (e.g. Fe 2+ , sulphide, and microbial metabolites; Armstrong, 1979; Armstrong and Armstrong, 2005; Mongon et al., 2014). Aerenchyma and the barrier to ROL in roots are regarded as key features contrib- uting to long-distance oxygen transport and waterlogging tolerance in many wetland species (Armstrong, 1979; Jackson and Drew, 1984; Justin and Armstrong, 1987; Colmer, 2003b; Colmer and Voesenek, 2009; Nishiuchi et al., 2012). However, the molecular mechanism of ROL barrier formation is poorly understood (Shiono et al., 2008).
 ==Plant Materials & Treatment==

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 ==The Background of This Project==
 <br>
 [[File: IC4R006-Microarray-2014-24913626-1.png|right|thumb|527px|'''Figure 3. Expression of selected suberin and lignin biosynthesis genes in the outer part of root (OPR) of rice.''']]
 <br>
 * The signal that triggers formation of the barrier to ROL is not known yet, but it may involve root exudates, cellular degradation products (Armstrong and Armstrong, 2001; Voesenek and Sasidharan, 2013) or phytotoxins present in chemically reduced waterlogged soils (e.g. Fe 2+ , sulphide, and microbial metabolites; Armstrong, 1979; Armstrong and Armstrong, 2005; Mongon et al., 2014). Aerenchyma and the barrier to ROL in roots are regarded as key features contrib- uting to long-distance oxygen transport and waterlogging tolerance in many wetland species (Armstrong, 1979; Jackson and Drew, 1984; Justin and Armstrong, 1987; Colmer, 2003b; Colmer and Voesenek, 2009; Nishiuchi et al., 2012). However, the molecular mechanism of ROL barrier formation is poorly understood (Shiono et al., 2008).
 [[File: IC4R006-Microarray-2014-24913626-2.png|center|thumb|727px|'''Figure 2. Numbers of up- and downregulated gene probes at the outer part of root (OPR) during formation of a radial oxygen loss (ROL) barrier in rice (i.e. the OPR along the basal part of long adventitious roots, 9 h treatment in deoxygenated stagnant 0.1% agar nutrient solution).''']]