2.1 The objectives of this chapter are to
2.3.2 Sodium and potassium content analysis
For sodium and potassium concentration measurement, the dried samples were pulverized, using a coffee grinder (KRUPS 75, Germany). One day before digestion the samples were dried overnight at 100 °C. 300 mg from each sample were placed in the microwave system MLS-MEGA II (Leutkirch, Germany). Four ml 65% HNO3 and two ml H2O2 30% (Roth, Germany) were added for each sample. The samples were placed in the microwave system at 200°C for 55 minutes under 15 atmospheric pressure (atm) and cooled down for 20 minutes. After digestion, the samples were diluted up to 25 ml using Seralpur water (deionized and filtered water). The samples were further diluted at 1:10 (0.5 ml plant material extract + 4.5 ml Seralpur water). The Na+ and K+ concentration was measured using the flame photometer Eppendorf, Elex 6361 (Hamburg, Germany). The flame photometer was calibrated along with ten samples using two calibration standards for both elements. The low standard was 0 mg/ l Na+ and 0 mg/ l K+ and the high standard was 100 mg/ l Na+ and 100 mg/ l K+. The Na+ and K+ contents were calculated as mg g-1 DW and the Na+/ K+ ratio was calculated.
2.3.3 Results
The salt treatment caused a significant reduction in the total plant FW and the total plant DW of the tested genotypes (Figures 3 and 4). The parental lines tested showed different interaction with salinity. The reductions in the entire plant FW were 60% for
20
cases of the Sollux and Gaoyou lines, the reductions in plant FW were 60% and 51%, respectively. Also the reductions in the total DW of Sollux and Gaoyou were 55% and 37%, respectively (Table 2, and Figures 3 and 4). The reductions in FW were 55%
and 60%, for Alesi and H30, respectively. The DW reductions were 39% and 40% for Alesi and H30, respectively. The biomass yield was higher in case of Mansholts, Samourai, Sollux, and Gaoyou lines compared with Alesi and H30. The variations between Alesi and H30 were non-significant (Table 3, and Figures 3 and 4). All parental lines showed a significant variation in the other parameters, such as the Na+ content of the 1st and the 2nd leaves and also the total plant Na+ content.
2.3.4 Conclusion
The flooding system is applicable and efficient. A number of parameters, including the FW, DW, Na+ content of the total plant and the Na+ content of 1st + 2nd leaves were relevant parameters to differentiate among the considered genotypes. As these parameters were suitable for phenotyping, the other parameters were not included in the next evaluation experiments. Because of the non-significant genotypic variation between Alesi and H30, they were not included in the further evaluation experiment.
0 2 4 6 8 10 12 14
Mansholts Samourai Sollux Gaoyou Alesi H30
Fresh weight (g)
Genotypes
Figure II-3: Total plant fresh weight (g) and signifcance levels of the six genotypes: Mansholts, Samourai, Sollux, Gaoyou, Alesi and H30
under control and salt stress Control
200 mM
**
**
* * ns
21
Chapter IIـــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــ
** Significant at P = 0.01; * significant at P = 0.05, + significant at P = 0.1 and ns = non-significant.
0 0.2 0.4 0.6 0.8 1 1.2 1.4
Mansholts Samourai Sollux Gaoyou Alesi H30
Dry weight (g)
Genotypes
Figure II-4: Total plant dry weight (g) mean values and signifcance levels of the six genotypes: Mansholts, Samourai, Sollux, Goyou, Alesi and H30 under control and salt stress
Control 200 mM
**
**
**
** ns ns
22
different parts in Mansholts and Samourai under control and salt stress (200 mM NaCl)
Genotypes Mansholts Mansholts Samourai Samourai
Traits / treatments control salt control salt
FW 1+2 (g) 2.50 2.36 1.94 1.21
FW 3+4 (g) 3.98 2.10 2.34 0.69
FW rest (g) 3.41 0.17 0.97 0.25
FW stem (g) 0.92 0.2 0.58 0.18
FW total Plant (g) 11.02 4.82 5.80 2.32
DW 1+2 (g) 0.21 0.26 0.17 0.12
DW 3+4 (g) 0.35 0.29 0.19 0.06
DW rest (g) 0.34 0.03 0.10 0.04
DW stem (g) 0.10 0.04 0.06 0.04
DW total Plant (g) 1.01 0.61 0.51 0.24
SPAD1 1+2 25.17 29.73 29.23 33.60
SPAD1 3+4 26.85 31.45 30.90 34.42
SPAD 2 1+2 24.85 35.28 29.52 34.38
SPAD 2 3+4 30.88 41.92 35.02 37.28
Na+ 1+2 (Na+ mg/ g DM) 3.83 16.40 0.97 15.79 Na+ 3+4 (Na+ mg/ g DM) 5.07 16.52 1.44 10.02 Na+ rest (Na+ mg/ g DM) 3.65 5.90 1.09 5.43 Na+ stem (Na+ mg/ g DM) 4.16 5.08 1.33 4.68 Na+ total Plant (Na+ mg/ g DM) 2.54 15.53 1.20 12.59 K+ 1+2 (K+ mg/ g DM) 20.98 6.61 12.39 11.80
K+ 3+4 (K+ mg/ g DM) 16.49 7.49 18.12 6.38
K+ rest (K+ mg/ g DM) 16.31 3.18 11.69 4.06 K+ stem (K+ mg/ g DM) 15.96 3.60 10.76 4.69 K+ Total Plant (mg K+ mg/ g DM) 17.41 6.80 14.26 9.23
23
Chapter IIـــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــ Table II-2: Experiment 2: The mean values of fresh weight (g), dry weight (g), SPAD values, sodium content Na+ mg/ g DM and potassium content K+ mg/ g DM of the different parts in Sollux and Gaoyou under control and salt stress (200 mM NaCl)
Genotypes Sollux Sollux Gaoyou Gaoyou
Traits / treatments control salt control salt
FW 1+2 (g) 4.93 1.90 3.44 1.88
FW 3+4 (g) 5.19 2.02 3.67 2.35
FW rest (g) 1.23 0.17 1.01 0.24
FW stem (g) 1.42 0.34 1.21 0.41
FW total Plant (g) 12.95 4.43 9.80 4.86
DW 1+2 (g) 0.41 0.21 0.35 0.25
DW 3+4 (g) 0.52 0.27 0.39 0.34
DW rest (g) 0.14 0.04 0.17 0.05
DW stem (g) 0.13 0.05 0.11 0.07
DW total Plant (g) 1.24 0.56 1.08 0.69
SPAD1 1+2 26.95 35.47 27.88 32.58
SPAD1 3+4 28.47 34.92 27.25 35.62
SPAD 2 1+2 28.40 40.10 30.17 38.62
SPAD 2 3+4 34.35 44.80 35.25 40.62
Na+ 1+2 (Na+ mg/ g DM) 1.38 14.95 4.31 17.69
Na+ 3+4 (Na+ mg/ g DM) 1.36 17.15 3.40 14.59
Na+ rest (Na+ mg/ g DM) 1.20 3.09 2.51 4.05
Na+ stem (Na+ mg/ g DM) 2.33 8.64 4.57 10.67
Na+ Total Plant (Na+ mg/ g DM) 1.46 14.39 3.43 15.23
K+ 1+2 (K+ mg/ g DM) 22.77 7.60 18.65 12.04
K+ 3+4 (K+ mg/ g DM) 17.10 9.60 15.72 9.29
K+ rest (K+ mg/ g DM) 14.24 3.09 15.68 4.65
K+ stem (K+ mg/ g DM) 23.89 5.19 17.42 6.59
K+ total Plant (K+ mg/ g DM) 19.50 7.95 17.73 9.90
24
different parts in Alesi and H30 under control and salt stress (200 mM NaCl)
Genotypes Alesi Alesi H30 H30
Traits / treatments control salt control salt
FW 1+2 (g) 2.35 1.67 2.47 1.27
FW 3+4 (g) 5.73 2.69 4.97 2.24
FW rest (g) 1.21 0.15 2.07 0.39
FW stem (g) 1.24 0.31 0.52 0.19
FW total Plant (g) 10.51 4.81 10.03 4.07
DW 1+2 (g) 0.20 0.17 0.21 0.16
DW 3+4 (g) 0.52 0.37 0.51 0.35
DW rest (g) 0.13 0.03 0.25 0.08
DW stem (g) 0.13 0.05 0.07 0.04
DW total Plant (g) 0.97 0.60 1.03 0.62
SPAD1 1+2 28.50 30.67 26.35 32.00
SPAD1 3+4 28.65 34.25 28.30 34.28
SPAD 2 1+2 29.25 35.97 26.35 35.75
SPAD 2 3+4 32.97 43.97 40.00 45.30
Na+ 1+2 (Na+ mg/ g DM) 2.59 10.60 2.84 12.98 Na+ 3+4 (Na+ mg/ g DM) 3.07 16.47 2.48 11.47 Na+ rest (Na+ mg/ g DM) 2.07 2.88 1.50 5.89 Na+ stem (Na+ mg/ g DM) 3.15 6.87 2.15 3.35 Na+ total Plant (Na+ mg/ g DM) 2.83 14.14 2.41 10.68 K+ 1+2 (K+ mg/ g DM) 19.70 8.50 20.19 12.75 K+ 3+4 (K+ mg/ g DM) 17.52 9.22 19.10 11.47 K+ rest (K+ mg/ g DM) 15.05 1.93 16.84 8.50 K+ stem (K+ mg/ g DM) 21.55 6.40 12.61 3.23 K+ total Plant (mg K+ mg/ g DM) 18.18 8.95 18.22 10.95
25
Chapter IIـــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــ 2.4 Experiment 3: Testing the performance of Mansholts × Samourai and Sollux
× Gaoyou under 200 mM NaCl 2.4.1 Material and Methods
The experiment was conducted in the greenhouse from the 23th of July until the 27th of August, 2012. The methods and calculations were the same as the previously mentioned protocol for Experiment 1 (2.2). Two weeks after beginning the 200 mM NaCl salt treatment, the plants were harvested and separated into 1st and 2nd leaves, and the remaining shoot system was labeled as “rest”.
2.4.2 Results
There was a significant reduction in the total plant FW and the total plant FW of the tested genotypes. The reductions in FW for Samourai and Mansholts were 62% and 53%, respectively. The reductions in DW for Samourai and Mansholts were 47% and 37%, respectively (Table 4). In case of, Sollux and Gaoyou lines, the respective reductions in FW were 66 % and 51%, and were 52% and 31% for the DW (Table 6, and Figures 5 and 6).
2.4.3 Conclusion
There was a significant difference between Mansholts and Samourai and between Sollux and Gaoyou, but the performance of both couples was similar (Figures 3 and 4). Nevertheless, the parental lines Mansholts and Samourai were considered to be suitable parental lines based on the molecular markers set and the quality of the linkage map.