International Journal of Agriculture and Crop Sciences.
Available online at www.ijagcs.com
IJACS/2012/4-15/1055-1059
ISSN 2227-670X ©2012 IJACS Journal
Effects of Different Levels of Salinity on Germination,
Proline Contents and A-, B- Chlorophylls in
Rapessed (Brassica napus L.)
Mehdi Hooshi Alavi1, Gholam Ali Ranjbar2
1. MSC of Plant Breeding, Sari Agricultural Sciences and Natural Resources University, Sari, Iran
2. Associate Professor, Sari Agricultural Sciences and Natural Resources University, Sari, Iran
Corresponding author email: [email protected],[email protected]
ABSTACT: Soil salinity is a major limitation for crop production in many areas of the world. Germination
and seedling responses of nine rapeseed cultivars (Hayola 401, Zarfam, Sarigol, Rgs003, L3006, Lph9,
Lph12, Lph22 and Lph23) to different levels of salinity stress (0, 100, 200 and 300 mM) evaluated in a
RCBD base on factorial experiment with three replicates in glasshouse. Results showed that the
germination of rapeseed cultivars were significantly affected by salinity. Cultivars Sarigol and Lph23
exhibited the highest and the lowest germination under salinity stress conditions. Amount of total
chlorophyll, chlorophyll a and b were significantly reduced by increasing salinity. The results showed that
increasing salinity levels significantly reduced chlorophyll a and b. Sarigol showed the most decrease in
the amount of total chlorophyll. Leaf and root proline content of cultivars have been significantly
increased by increasing the NaCl level, so that, the most increasing was happened using 300 mM NaCl
treatments.
Keywords: Rapeseed, salinity stress, germination, chlorophyll and praline
INTRODUCTION
Salinity is one of the major abiotic stresses in arid and semi-arid regions that substantially reduce the
average yield of major crops by more than 50% (Bray, 2000). Salinity affects 7% of the world's land area for
around 930 million ha (Munns, 2006). Every year more and more lands become non-productive due to salt
accumulation. Therefore, understanding the mechanisms of plant tolerance to salinity stress is important (Bartels
et al., 2005). Although salt stress affects all growth stages of a plant, seed germination and seedling growth stages
are known to be more sensitive in most plant species (Cuartero, 2006). Furthermore, germination and seedling
stage is predictive of plant growth responses to salinity (Blum, 1985). Therefore, seeds with more rapid
germination under salt stress and/or normal conditions may be expected to achieve a rapid seedling establishment
and more salt tolerance, resulting in good stand establishment and hence higher yields (Munns, 2002). Successful
seedling establishment depends on the frequency and the amount of precipitation as well as on the ability of the
seed species to germinate and grow while soil moisture and osmotic potentials decrease (Roundy, 1985).
Moreover, salt stress has also been found responsible for an increased respiration rate, ion toxicity (Sudhir
et al., 2004), decreased biosynthesis of chlorophyll (Khan et al., 2009) and inefficiency of photosynthesis (Munns,
2002), all of which ultimately leading to lowered economic productivity. One of the effects of salinity stress is
reducing photosynthetic activity which caused to decreasing b and a chlorophylls and reducing Co2 uptake and
photosynthetic capacity (Francois et al., 1993). Proline is used as an enzymic protector that contributes in
macromolecules structure and is main source of energy and nitrogen to confront salinity. According to studies,
Iqbal et al (2006) reported that chlorophyll is reduced under saline conditions. Nazarbeygi et al (2011) reported
that with increasing salinity, reduction in the rate of chlorophyll and increase in proline in various rapeseed cultivars
were significantly different. Rapeseed is an important oil crop often cultivated in arid and semiarid regions of the
world such as Iran where salinity threatens to become, or already is, a problem. Present study was conducted to
consider effects of salinity on seed germination, seedling growth, Proline and a-, b- Chlorophylls of nine rapeseed
genotypes cultured under salinity conditions.
Intl J Agri Crop Sci. Vol., 4 (15), 1055-1059, 2012
MATERIALS AND METHODS
Germination experiment
Nine rapeseed cultivars (Brassica napus L. c.v's Hayola 401, Zarfam, Sarigol, Rgs003, L3006, Lph9,
Lph12, Lph22 and Lph23) were collected from Agricultural Research Center of Sari, Iran. The same size seeds
from each cultivar were surface-sterilized for 5 min in sodium hypochloride solution (10%) and then rinsed with
distilled water for 3 to 5 times. After sterilization, 25 seeds were transferred into 9 cm sterile petri dishes on filter
paper and then were wetted with 7 ml distilled water (control) or saline water solution at 0, 100, 200 and 300 mM
NaCl levels. To prevent infection and evaporation of solution, all plates were sealed using parafilm and culture
operations were performed under a laminar air flow cabinet in sterile conditions. The Petri dishes were labeled and
incubated in a germinator at 25 °C and 12/12 h day/night photoperiod. Measurement of germinated seed was done
daily until end of seventh day. At the end of germination, root length were tested and shoot and root and shoot
fresh weight were measured. The germination percentage (GP) and final germination percentage (FGP) were
calculated for all cultivars as follow:
Where, n: Number of germinated seed at seventh day; N: number of seeds
Where, n: Number of germinated seed at lightly day; t: days after beginning
Glasshouse experiment
Ten seeds of each cultivar were sown directly in plastic pots containing 4 kg of field soil. The bottom of
each pot was delved for drainage of extra water. Pots were transferred to glasshouse under conditions of 25/18 °C
day/night temperature and natural light. After full germination, the number of plants was reduced to three seedlings
per pot. Salinity stress induction was done when fourth leaf was completely expanded. Different concentrations of
NaCl solution was added to each pot for 15 days. In order to assess leaf chlorophyll, leaf samples were harvested
and dried for 48 hours at a temperature of 75 °C. Hiscox method (1979) was used to evaluate the amount of
chlorophyll and Lichtenthaler equation (1987) was exploited to calculate them numerically. For measuring Proline
content, Bates metod (1973).was used. The experiment was carried out using a factorial experiment based on
randomized block design with three replications. Analysis of variance (ANOVA) was carried out using general
linear model (GLM) in SAS (6.12 version) statistical software. Also, for comparison of means Duncan's Multiple
Range Test (DMRT) was used.
RESULTS AND DISCUSSION
The results of analysis of variance related to the percentage, germination speed, root length and shoot
lenght showed that these traits were influenced by genotype (Table 1). The interaction of salinity cultivar on
germination rate and germination speed significant and root length and shoot lenght non-significant. These figures
indicate that the effect of different concentrations of salinity in terms of response rate and percentage germination
were different. Mean comparisons of different canola genotypes showed that Varieties Sarigol and Lph23 highest
and lowest germination under salt stress conditions, respectively. Fast decreasing of germination of genotypes
Lph22and Lph12 by increasing salinity showed sensitivity of these genotypes to salinity. (Table 2). These results
are in agreement with research of Demir (2003). Increasing salinity levels, reduced germination rate and reduced
final germination percentage was followed (fig 1 and 2). Results showed the negative effects of salinity on root and
shoot length (fig 3 and 4). In addition, according to the results of Werner and Finkelstein (1995) salinity decreases
water absorption and growth of root and shoot.
The results of analysis of variance related to prolin, clorophyll a and b showed that Interaction of cultivar ×
salinity on proline, chlorophyll a and b in different amounts of rapeseed cultivars and salinity levels of chlorophyll a,
chlorophyll b, chlorophyll a to b ratio and total chlorophyll was significant (Table 3). Results showed that amount of
total chlorophyll, chlorophyll a and b significantly reduced by increasing salinity levels (Table 4). Cultivar Sarigol
showed the highest decrease in total chlorophyll than the other genotypes. The decrease in chlorophyll contents
Intl J Agri Crop Sci. Vol., 4 (15), 1055-1059, 2012
under saline conditions was reported by Ashraf et al (2005). Also, results indicated that proline content of leaf and
root has significantly increased due to increasing in NaCl rate. So that, treatment 300 mM NaCl demonstrated
more proline content than the other treatments (fig 5). Valia et al (1993) suggested that increasing of proline
content under saline conditions caused less existing glutamate in biosynthesizing chlorophyll, thus, it helped to
producing proline for plant bearing more salinity conditions.
Table 1. Analysis of variance on germination traits under salinity stress
S. O. V
DF
MS
Germination speed
Repeat
Salinity
Cultivar
Cultivar Salinity
Eror
2
3
8
24
69
2.79
**
26.31
**
6.10
**
0.72
0.31
Germination percentage
**
**
Shoot length (cm)
**
2836.93
**
19962.70
**
3860.13
*
555.68
377.17
Root length (cm)
**
0.7
**
14.15
**
0.43
ns
9.57
**
140.50
**
2.71
1.38
0.06
ns
1.46
1.02
*, ** significant at the 0.05 and 0.01 probability levels, respectively. ns, not significan.
Table 2. Comparison of main effect of cultivars on germination traits
Cultivar
Germination speed
Hayola401
Lph9
Lph12
Lph22
LpH23
RGS003
L3066
Zarfam
Sarigol
ab
Mean
Germination percentage
Shoot length (cm)
a
2.66
b
2.29
c
1.64
c
1.68
d
0.63
ab
2.53
ab
2.68
b
2.24
a
2.89
Root length (cm)
a
76.00
ab
66.00
bc
52.67
c
49.33
d
21.08
a
72.67
a
73.00
abc
63.67
a
76.42
abc
1.22
ab
1.00
b
0.98
ab
1.00
c
0.53
b
0.98
ab
1.04
ab
1.04
ab
1.10
2.34
c
1.72
a
3.02
abc
2.26
bc
1.75
a
2.98
abc
2.64
abc
2.65
ab
2.67
For a given means within each colum followed by the same letter are not significantly differences (P < 0.01).
Table3. Analysis of variance on prolin, chlorophyll a and b under salinity stress
S. O. V
DF
Repeat
2
Salinity
Cultivar
Cultivar Salinity
Eror
8
3
24
69
MS
Prolin
128.13
Chla
**
2413.32
**
403.88
**
52.75
2.32
2.27
**
Chlb
ns
12.65
**
698.54
**
107.25
**
12.28
2.57
Chla+Chlb
*
4.22
**
797.35
**
125.58
**
23.41
4.09
ns
2960.44
**
406.38
**
46.84
5.38
Chla/Chlb
1.13
**
**
**
6.70
**
2.38
**
0.88
0.2
*, ** significant at the 0.05 and 0.01 probability levels,respectively. ns, not significan.
Table 4 . Mean comparisons of chlorophyll content of rapeseed treated with NaCl
NaCl
treatment
(mM)
0
100
200
300
Mean
Chla
Chlb
a
22.31
b
19.57
c
14.45
d
10.86
Chla+Chlb
a
17.28
b
11.91
c
8.10
d
4.47
a
39.59
b
31.48
c
22.55
d
15.33
Chla/Chlb
c
1.47
b
1.81
b
2.02
a
2.67
For a given means within each colum followed by the same letter are not significantly differences (P < 0.01).
Intl J Agri Crop Sci.
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1055-1059
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Figure 1 . Effects of different salinity levels on germination
germination rate of rapeseed cultivars.
Figure 2 . Effects of different salinity levels on germination percentage of rapeseed cultivars.
Figure 3 . Effects of different salinity levels on radicle length of rapeseed cultivars.
cultivars
Figure 4 . Effect of different salinity levels on plumule length of rapeseed cultivars.
cultivars
Intl J Agri Crop Sci.
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(
1055-1059
1059, 2012
Figure 5 . Proline content changes in rapeseed cultivars under different NaCl concentrations.
concentrations
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