Impact of Salt Stress (NaCl) on Seed Germination, Photosynthetic Pigments of Green Gram Cultivars of Co6 and Co8  

K. Krishna Surendar , S.V. Varshini , R. Deepa Sankari , N. Susithra , S. Kavitha , M. Shankar
Vanavarayar Institute of Agriculture, VIA, Pollachi-642 103, India
Author    Correspondence author
Plant Gene and Trait, 2014, Vol. 5, No. 6   doi: 10.5376/pgt.2014.05.0006
Received: 22 May, 2014    Accepted: 30 May, 2014    Published: 13 Jun., 2014
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Surendar et al., 2014, Impact of Salt Stress (Nacl) on Seed Germination, Photosynthetic Pigments of Green Gram Cultivars of Co6 and Co8, Plant Gene and Trait, Vol.5, No.6 40-44 (doi: 10.5376/pgt.2014.05.0006)

Abstract

Investigations were undertaken to study the impact of salt stress (NaCl) in concentrations on seed germination and seedling growth of Green gram (CO5, CO6). Seed germination percentage, seedling growth characters, physiological and bio-chemical parameters were estimated at 10 days after sowing in Petridish. The stress was imposed during sowing time with different concentrations viz., 10, 20, 30, 40, 50, 60, 70, 80, 90, 100 ppm. The increased seed germination percentage was noticed in control (distilled water) treated seeds and very less reduction was observed in T2 to T5 treatments in the range of 11.2 percent over the T11. The highest reduction percent of 35% were observed in T10, T11, & T12 treated seeds. Growth parameters of shoot length and root length were significantly reduced due to NaCl treatments. However among the treatments, T1- T5 showed very less reduction 15 percent than the other treatments, whereas T6-T11 recorded highest reduction of 27 percent over control. There was also significantly maintained in the chlorophyll ‘a’ ‘b’ and total chlorophyll content of the seedlings in the T1-T5 treatments as 12.6 %, with lesser reduction over the other treatments. The highest reduction of 25.3 percent was noticed in T6-T12 treated seedlings.

Keywords
Salt stress; Seed germination; Shoot and root length; Photosynthetic pigments; Green gram

Legumes are the richest protein source of human diet and livestock in poor areas. Apart from that, they are used as green manures and green fodder to animals. Mainly they are used for fixing atmospheric nitrogen to improve the physical and chemical properties of soil. Among the legumes, Green gram and Black gram are considered as the most important traditional crops of India.Salinity – an abiotic stress is an ever increasing problem that seriously affects crop production in various parts of the world, especially in areas where are irrigated with water containing salts. About 23% of the world’s cultivated lands are saline and 37% is sodic (Khan and Duke, 2001). Salinity affects 7% of the world’s land area of about 930 million hectares. Salinity reduces the yield of pulses by more than 50% (Bray, 2000). Soils can be saline due to geo-historical processes or they can be man-made. The water and salt balance, just like in oceans and seas determine the formation of salty soils, where more salt comes in than goes out. Here, the incoming water from the land brings salts that remain because there is no outlet and the evaporation water does not contain salts. Soil salinity in agriculture soils refers to the presence of high concentration of soluble salts in the soil moisture of the root zone. Salt stress induces the synthesis of abscisic acid which closes stomata when transported to guard cells. As a result of stomatal closure, photosynthesis declines and photo inhibition and oxidative stress occur. The deleterious effect of salinity is increased osmotic pressure which restricts the absorption of water into the seeds (Tester and Davenport, 2003). It is also toxic to the embryo and seedlings. Enzyme called α-amylase which is essential for seed germination is inhibited due to salt stress. Starch to sugar conversion occurs during germination is also affected by salinity. It also delays the synthesis of nucleic acids and RNAase. As regard to the chlorophyll content of the salinized plant, it is apparent that the chlorophyll content was reduced with increasing salinity. When salinity has affected the warning signs were sick or dying trees and declining vegetation. As salinity impacts on any remaining native vegetation and the wildlife that depends on it for survival, the loss of biodiversity escalates. Salinity also reduces the productivity of crops and the sustainability of agriculture. Based on the above constraints, we are taken the following objectives,
Screening of Green gram varieties for NaCl stress tolerance through morphological analysis.
To determine the physiological basis of NaCl stress tolerance in Green gram varieties.
1 Materials and Methods
The experiment was carried out at Vanavarayar Institute of Agriculture (TNAU affiliated), Pollachi, Tamil Nadu, India during 2013-2014. The experiment consists of ten treatments with three replications were laid out in completely randomized block design with two cultivars of CO5 and CO6. Seeds of green gram varieties obtained from Department of Pulses, Tamil Nadu Agricultural University, Coimbatore, were used for the study and the details of the varietal characters were listed in Table 1. Green gram varieties (Table 1) were screened for tolerance to various levels of sodicity stress, based on germination per cent, seedling growth and vigour index, seeds were allowed to germinate in Petri dishes. The germination medium was prepared following the procedure mentioned below. Petri dishes were sterilized using 0.01% HgCl2 and 70% ethanol and finally washed with distilled water. Before placing the germination sheet, Petri dishes were cleaned thoroughly with a cotton swab. The surface sterilized (70% ethanol) 15 seeds from each variety were placed in each Petri dish. For imposing sodicity (11 levels as considered as Treatments) stresses, sodium chloride (NaCl) solution at the concentration of T1: control (without NaCl), T2:10, T3:20, T4:30, T5:40, T6:50, T7:60, T8:70, T9:80, T10:90 and T11:100 ppm were prepared. The seeds were allowed to germinate, by sprinkling the salt solution of 10 ml each in alternate days. Distilled water was used for maintaining the control. The pH and EC details of the salt solution used for experiment were given in Table 2.


Table 1 Varietal details



Table 2 pH and EC of the salt solution used for experiment


1.1 Observation recorded
The germination percentage, root length, shoot length and Vigour index were measured at 15th DAS and chlorophyll ‘a’ ‘b’ and total chlorophyll content was estimated based on the procedure given by Yoshida et al. (1979) and expressed as mg/g fresh weight.
1.2 Germination percentage
The germinability was recorded on the fifteenth day after sowing (DAS) and number of seeds germinated was expressed as per cent.



1.3 Seedling shoot length
On 15th DAS, seedlings from each replication were carefully removed at random. Length of shoot was measured from the collar region to the tip of the longest leaf and expressed as cm.
1.4 Seedling root length
Root length of the seedlings was measured from the base of the stem to the tip of the longest root and expressed as cm.
1.5 Vigour index
The vigour index of the seedlings was calculated using the following formula proposed by Abdul-Baki and Anderson (1973) and expressed as per cent.
VigourIndex=(Shoot length+Root length)×Germi- nation percentage
1.6 Photosynthetic pigments                                                   
The chlorophyll content was estimated following the method suggested by Yoshida et al. (1971) and expressed as mg g-1 fresh weight.
1.7 Procedure
Take 250 mg of leaf sample is macerated with 10ml of 80% acetone using a pestle and mortar and the extract is centrifuged at 3000 rpm for 10 minutes. The supernatant solution is transferred into a 25ml volumetric flask and made up to 25ml using 80% acetone. Then the color intensity of the green pigment is read at 480nm, 510nm, 645nm, 663nm and 652nm for chlorophyll a, chlorophyll b and total chlorophyll content respectively using Spectrophotometer.
1.8 Calculation



Total Chlorophyll=Chlorophyll a+Chlorophyll b

2 Result and Discussion
2.1 Germination percentage
The result on seed germination percentage was significantly differed in all the treatments. Among the treatments, T1 showed highest germination percentage in green gram both CO6 and CO8 (99.7 and 99.7), which was followed by T2, T3, T4 and T5. The lowest germination percentage was recorded in T9, T10 and T11 treatments. The treatments of T9 and T11 in CO6 and T2 and T3, T7 and T8 were on par with each other (Table 3).


Table 3 Effect of salt stress (NaCl) on germination percentage, shoot length, root length and vigour index of green gram


The increased seed germination percentage was noticed in control (distilled water) treated seeds and very less reduction was observed in T2 to T5 treatments in the range of 11.2 percent over the T11. The highest reduction percent of 35% were observed in T10, T11, and T12 treated seeds. Enzyme called α – amylase which is essential for seed germination is inhibited due to salt stress. Starch to sugar conversion occurs during germination is also affected by salinity. It also delays the synthesis of nucleic acids and RNAase (Tester and Davenport, 2003).
2.2 Shoot length (cm) and Root length (cm)
The result on shoot length was significantly differed in all the treatments. Among the treatments, T2 showed the highest shoot length in green gram both CO6 and CO8 (9.6 and 10.32), which was followed by T1, T3, and T4. The lowest shoot length was recorded in T10, and T11 treatments (Table 3).
The result on root length was significantly differed in all the treatments. Among the treatments, T1 showed highest root length in green gram both CO6 and CO8 (5.92 and 5.90), which was followed by T2, and T3. The lowest root length was recorded in T9, T10 and T11 treatments. The treatments of T9 and T10 in CO6 and CO8 were on par with each other.
Growth parameters of shoot length and root length were significantly reduced due to Nacl treatments. However among the treatments, T1~T5 showed very less reduction 15% than the other treatments, whereas T6-T11 recorded highest reduction of 27% over control. Length of root and shoot decreased perhaps due to accumulation of ions near the root surface (Hanson, 1978).
2.3 Vigourindex
The result on Vigour index was significantly differed in all the treatments. Among the treatments, T1 showed highest Vigour index in green gram both CO6 and CO8 (1460.85 and 1584.43), which was followed by T2, T3, and T4. The lowest Vigour index was recorded in T10 and T11 treatments (Table 3).
2.4 Chlorophyll a (mg/g)
The result on Chlorophyll ‘a’ was significantly differed in all the treatments. Among the treatments, T1 showed highest Chlorophyll ‘a’ content in green gram both CO6 and CO8 (3.35 and 1.86), which was followed by T2, T3, T4, T5 and T6. The lowest Chlorophyll ‘a’ content was recorded in T10 and T11 treatments (Table 4).


Table 4 Effect of salt stress (NaCl) on chlorophyll a, chlorophyll b and total chlorophyll content of green gram


2.5 Chlorophyll b (mg/g)
The result on Chlorophyll ‘b’ was significantly differed in all the treatments. Among the treatments, T1 & T2 showed highest Chlorophyll ‘b’ content in green gram both CO6 and CO8 (0.78and 0.32), which was followed by T3, T4, T5 and T6. The lowest Chlorophyll ‘b’ content was recorded in T10 and T11 treatments (Table 4).
2.6 Total Chlorophyll (mg/g)
The result on total chlorophyll content was significantly differed in all the treatments. Among the treatments, T1 showed highest total chlorophyll content in green gram both CO6 and CO8 (4.41 and 2.95), which was followed by T2, T3, T4 and T5. The lowest total chlorophyll content was recorded in T9, T11 and T10 treatments (Table 4).
There was also significantly maintained in the total chlorophyll content of the seedlings in the T1~T5 treatments as 12.6%, with lesser reduction over the other treatments. The highest reduction of 25.3 percent was noticed in T6~T12 treated seedlings. According to Lapina and Popov (1970), saline conditions lead to disruption of the fine structure of chlorophyll and instability of the pigment protein complex which may also be the cause of the reduced chlorophyll content.
References
Abdual-baki A.A., and J.D. Anderson, 1973, Relationship between decarboxylation of glutamic acid and vigour in soybean seed, Crop Sci., 13: 222-226
Bray E.A., J. Bailey-Serres, and E. Weretilnyk, 2000, Responses to abiotic stress, pp. 1158-1203. In: Buchanan B., Gruissem W. and Jones R. (eds.), Biochemistry and Molecular Biology of Plants. American Society of Plant Physiology. , Rockville
Hanson, 1978. Application of the chemiosmotic hypothesis to ion transport across the root, Plant Physiol., 42: 294-298
Khan M.A, and Duke N.C., 2001, Halophytes-A resource for the future, Wetland Ecology and Management, 6:455-456
http://dx.doi.org/10.1023/A:1012211726748
Lapina I.P., and Popov, 1970. Effect of sodium chloride on the photosynthetic apparatus of tomatoes, Fiziol. Rast., 17: 580-585
Tester M., and Davenport R., 2003. Na+ tolerant and Na+ transport in higher plants, Annals of Botany, 91: 503-527
http://dx.doi.org/10.1093/aob/mcg058

Yoshida S., Forno D.A, and Cock J.H., 1976, Gomez KA, Laboratory Physiological Studies of Rice, Phillipines, Lagena: Int. Rice Res. Inst. (IRRI)

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