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The Effects of Salinity on The Germination, Root Height, Stem and Plant Height

Abstract

Soil salinity is one of the most important constraints limiting crop production in arid and semi arid regions. Seed germination is a critical stage in the life history of individual plants and salt tolerance during germination is crucial for the establishment of plants growing in saline soils. The research was carried out to find out the effect of salinity on the germination, root height, and stem height and plantlet height of three local varieties. Five different concentration of saline water was made from 0 to 8g/l. The plant was provided with this saline water. It was noted that rate of germination, root height, stem height and plantlet height decreased with the increased in salt concentration. Highest germination percentages and other growth parameters related to control (0g/L) in all varieties and lowest germination occurs at 8g/L

Introduction

1.0. Overview:

The Brassicaceae is a family of plants that provide food, fodder and forage (Dixon, 2007). Increased interest in energy self sustainability in the United States has brought new attention to the importance of the Brassicaceae including Brassica napus, mustard (Brassica juncea) and camelin Camelina sativa (Putnam et al., 1993) as biofuel feedstocks. In the United States production of Brassicas for biofuel has until recently been primarily in the northern tier of states. In that portion of the US, and adjacent areas of Canada, the concern is how to best germinate seed at temperatures from 10 to 22 ◦C (Nykiforuk and Johnson-Flanagan, 1994; Zheng et al., 1994; Vigil et al., 1997; Willenborg et al., 2004). Brassica production is being considered for other geographic areas in the United States where higher soil temperatures could occur at sowing. Varieties of winter hardy Brassica napus have been developed for use in the southern Great Plains with establishment dates from mid-August to late-September for harvest the following spring (Boyle et al., 2004a, and b). Soil temperatures range from 23 to 28 ◦C. Russo and Bruton (2008) reported that planting dates for Brassica napus in Oklahoma could be extended to late-October for some varieties; soil temperatures can be 20 ◦C at this time.

Brassica juncea, also known as Indian mustard or mustard greens or leaf mustard, is perennial herb, usually grown as an annual or biennial.

The English word “mustard” is derived from the Middle English “moustarde”, a combination of the Old French words “moust” which means must and “ardens” meaning burning (Antol, 1999). “Moust” is derived from the Latin “mustum”, meaning “new wine”. Romans were the first to experiment with the preparation of mustard as a condiment. They mixed unfermented grape juice, known as “must” with ground mustard seeds to make “burning must”, mustum ardens-hence “mustard” (Hazen, 1993).

Brassica juncea is a herbaceous plant with an erect, branched stem up to 1.0 m tall, with a taproot reaching 60-80 cm in depth, lower leaves petioled, green, sometimes with a whitish bloom, ovate to obovate, variously lobed with toothed or frilled edges; upper leaves subentire, short and petioled, constricted at intervals, sessile. The flowers consist of 4 (four) yellow petals arranged in a cruciform manner, 4 (four) yellowish green sepals, a short green pistil with a knobby stigma, and tetradynamous stamens with yellow anthers. They are pollinated by bees that soon develop into sickle-shaped green seed pods. Seeds are sown in very early spring. Plants are generally harvested before fruits are fully ripe to reduce shattering.

The growing period is from 40–60 days depending on the variety and weather conditions. Indian mustard is a cool-season vegetable, growing well at monthly average temperatures ranging from 15 to 18°C. It can tolerate annual precipitation of about 500 to 4,200 mm, annual temperature of 6 to 27°C and pH of 4.3 to 8.3.

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1.1. Origin and Distribution:

The primary center of origin of mustard is thought to be central Asia while secondary centers in central and western China. Major mustard producing countries include Canada, China, Germany, France, Australia, Pakistan, Poland and India. (Duhoon et al., 1998; Dutta et al., 2008; Misra et al., 2010)

1.2. Economic importance:

 Brassica juncea is valued for its intense flavours and healing properties. This plant is cultivated mainly as an oil crop. It is a good bee plant (Fomina, 1962). All over the world, mustard is used for its appetizing flavor and preservative value and the seeds are used largely for tempering food. Mustard is available in the form of seeds, powders and oil. Recently, Brassica juncea has been explored for its bio diesel potential (Jham et al., 2009). Its importance is listed below:

1.3. Nutritional importance:

The plant appears in some form in African, Indian, Chinese, Japanese, and Soul food cuisine. The leaves, the seeds, and the stem of Indian mustard are edible. Seeds of Brassica juncea contain 25-30% fatty non-drying oil and glycoside sinigrine. The leaves of young plants are used as green vegetables as they supply enough sulphur and minerals in the diet. Brassica juncea is used to make the pickle called Achar, and the Chinese pickle known as „Zha cai‟ (Everitt, 2007). Young tender leaves of mustard greens are used in salads or mixed with other salad greens. Older leaves with stems may be eaten fresh, canned or frozen, for potherbs, and to a limited extent in salads. Its basal leaves are eaten raw and used in salads or cooked like spinach. Leaves and stems are also added to soups and stews. Mustard greens are often cooked with ham or salt pork, and may be used in soups and stews. Seed residue is used as cattle feed and in fertilizers. In Asia, some kinds of mustard are pickled. (Grubben and Denton, 2004).

The seeds are very pungent and used to season meats and other dishes. Although widely and extensively grown as a vegetable, it is being grown more for its seeds which yield an essential oil and condiment. Oil is used to pickle foods in Kashimiri and Bengali cooking. It is used as cooking oil in parts of India and Bangladesh. Mustard oil is one of the healthiest edible oils. Mustard oil is healthier than olive oil because it has no fats, has low saturated fats, high mono-unsaturated fats and polyunsaturated fatty acids such as omega-3. It is stable at high temperatures, which makes it ideal for Indian cooking and even deep frying. Mustard is also a cheaper alternative of edible oil and makes the food tastier. In very small amounts, it is often used by the food industry for flavoring. Mustard seed meal is good source of protein (28−36%) and phenolic antioxidants such as sinapine and sinapic acid (Das et al., 2009). Mustard oil is also used as hair oil, lubricants and as a substitute for olive oil in Russia. Oilseed cake is used for cattle feed and manure.

1.4. Medicinal value:

 Seeds are used for curing tumors in China whereas roots are used as a galactagogue in Africa. Ingestion may impart a body odor repellent against mosquitoes (Burkill, 1966). Believed to be aperients and tonic, the oil is used as a counter-irritant and stimulant. In Java, the plant is used as an ant syphilitic emmenagogue. Leaves applied to the forehead are known to relieve headache (Burkill, 1966). Chinese eat the leaves in soups for bladder inflammation or hemorrhage. Mustard oil is used for skin eruptions and ulcers (Perry, 1980). The seeds, crushed in honey, are known to cure coryza. Swallowing mustard seeds soaked in mustard oil, cures stomachache. Massaging the body with mustard oil is very beneficial as it cures flatulence and makes the body strong. Massage with the oil is thought to improve blood circulation, muscular development and good texture to human skin. The oil is also antibacterial. In skin diseases, the local application of seed oil is beneficial as it is antiseptic and anti-inflammatory. Derivatives of the mustard constituent i.e. allyl isothiocyanate, forms the basis for toxic agents such as mustard gases of warfare and the antineoplastic nitrogen mustard. Mustard also has antioxidant activity and pharmacological effects on cardiovascular disease, cancer and diabetes. The dried, ripe seeds are used commercially. Mustard and its oil have been used as a topical treatment for rheumatisms and arthritis. Mustard seeds have been used as appetite stimulants, emetics, and diuretics. The oil has a strong smell, a hot nutty taste, and is mainly used for cooking in Bengal, Bihar and other areas of India and Bangladesh. The oil makes up about 30% of the mustard seeds. Mustard oil is beneficial to human health because of its low content of saturated fats, ideal ratio of omega-3 and omega 6 fatty acids, content of antioxidants such as vitamin E and also of the fact that it is cold pressed (extracted at 45-500C). The pungent taste of the mustard condiment results when ground mustard seeds are mixed with water, vinegar, or other liquids (or chewed). Under these conditions, a chemical reaction between the enzyme myrosinase and a glucosinolate known as sinigrin from the seeds leads to the production of allyl isothiocyanate. The main component of mustard oil is allyl isothiocyanate (Yu et al., 2003). Allyl isothiocyanate serves the plant as a defence against herbivores. Since it is harmful to the plant itself, it is stored in the harmless form of a glucosinolate, separate from the myrosinase enzyme. Once the herbivore chews the plant, allyl isothiocyanate is produced. Seeds of this plant are widely used in America, Japan, China and other countries and regions as a traditional pungent spice, a source of edible oil and protein and a kind of medicine. Brassica juncea, an amphidiploids species, is grown as an oilseed crop in India. Although, oilseed Brassicas are grown over 15% of arable land in India but their productivity is considerably hindered by various biotic and abiotic stresses (Shah, 2002; Purty et al., 2008; Sirhindi et al., 2009; Yusuf et al., 2010), like drought, chilling, pesticides and heavy metals etc. The soil in which plants grow may contain phytotoxic levels of the heavy metals including Cr, Cu, Hg and Ni, Zn etc.

The knowledge of seed biology is essential for understanding the process and patterns within a plant community, such as the establishment of plants, seed development and germination (Godoi and Takaki 2004). It is widely accepted that successful seedling establishment depends on a seed’s ability to germinate (Baskin 2005). Stress and strain are fundamental physical concepts that can be applied to biological systems. Physical scientists define stress as a force per unit area applied to an object. Strain is a change in a dimension of an object developed in response to a stress (Hopkins and Huner 2003). Salinity is an increasing environmental problem worldwide. Salts in the soil water may inhibit plant growth for two reasons. First, the presence of salt in the soil solution reduces the ability of the plant to take up water, and this leads to reductions in the growth rate. This is referred to as the osmotic or water-deficit effect of salinity. Second, if excessive amounts of salt enter the plant in the transpiration stream there will be injury to cells in the transpiring leaves and this may cause further reductions in growth. This is called the salt-specific or ion-excess effect of salinity (Yao and Fang 2009). The definition of salt tolerance is usually the percent biomass production in saline soil relative to plants in non-saline soil, after growth for an extended period of time. For slow-growing, long-lived, or uncultivated species it is often difficult to assess the reduction in biomass production, so percent survival is often used. The yellow mustard (Brassica juncea) is an important oilyielding crop of saline lands, but information on salt tolerance, which indicates its salt-prone nature at the germinating stage (Kumar 1984).

Each plant species has a specific range of environmental requirements necessary for germination (Baskin and Baskin, 2001). Seed germination is a complex biological process that is influenced by various environmental such as temperature, light, pH and soil moisture, and genetic factors (Shafii and Price, 2001; Chachalis and Reddy, 2000). Temperature is a major environmental factor that affects the seed germination capacity and germination rate (Kamkar et al., 2012, Kebreab and Murdoch, 2000). Temperature has an impact on a number of processes that regulate seed germination, including membrane permeability and the activity of membrane-bound and cytosolic enzymes (Tlig et al., 2008). Germination rate usually increases linearly with increasing temperature up to an optimum point, and then decreases linearly to a ceiling temperature (Steinmaus et al., 2000; Bradford, 2002). Optimum temperature is a temperature that seed showed highest germination in minimum time (Alvarado and Bradford, 2002).

Too, germination of many plant was affected by water potential of soil that simulated by poly ethylene glycol (Rashed-Mohassel et al., 2012; Biligetu et al., 2011; Kazerooni Monfared et al., 2012). Seeds must reach a critical water content to trigger cell elongation and initiate radicle emergence (Bradford, 1995). Seed germination rates generally decrease with decreases in water potential, which is always associated with failure of plant emergence (Willenborg et al., 2005). In addition, water and temperature often interact in regulating seed germination. For example, seeds are capable of germinating at higher levels of water stress at optimum temperatures (Kebreab and Murdoch, 1999).

Different levels of heavy metals contained in soil, water and air cause pollution after reaching certain concentrations. There are different reasons for this kind of pollution. Metals are continuously released into the biosphere by volcanoes, natural weathering of rocks and by industrial activities such as mining and the combustion of fossil fuels and the release of sewage (De Abreu et al., 1998). Heavy metal contamination of agricultural soil has also been observed to be increased due to industrialization. Therefore, heavy metal contamination represents a risk for primary and secondary consumers and ultimately humans (Zeller and Feller, 1999).

Agricultural soils, as an essential part of the environment, are no exception of this phenomenon. Cadmium (Cd), copper (Cu), lead (Pb), and zinc (Zn) are among the most abundant heavy metals in the agricultural soils (Förstner, 1995). These metals are mostly absorbed by plants easily and prove toxic to plants that can be observed as growth retardation as a result of alterations in biochemical process like inhibition of enzyme activity, protein penetration and impaired nutrition etc. (Arun rtal, 2005). In the recent era, lead (Pb) contamination has gained a considerable attention as a potent environmental pollutant. Significant increases in the Pb content of cultivated soils have been observed near urban and industrial areas where it tends to accumulate in the surface ground layer (Di Toppi and Gabrielli, 1999). Despite regulatory measures adopted in many countries to have a check on Pb input in the environment, it continues to be one of the most serious global environmental hazards in the developing world (Yang et al., 2000).

Cadmium (Cd), being a highly toxic metal pollutant of soils, inhibits root and shoot growth and yield production, affects nutrient uptake and homeostasis, and is  frequently accumulated by agriculturally important crops and then enters the food chain with a significant potential to impair animal and human health (Di Toppi and Gabrielli, 1999). The reduction of biomass by Cd toxicity could be the direct consequence of the inhibition of chlorophyll synthesis and photosynthesis (Padmaja, 1990). Excessive amount of Cd may cause decreased uptake of nutrient elements, inhibition of various enzyme activities, induction of oxidative stress including alterations in enzymes of the antioxidant defence system (Sandalio et al., 2001)

Aims and objectives:

  • To study the germination response of mustard plant of different regions.
  • To find out the best conditions for mustard plant germination.
  • To study the effect of salinity on mustard plant.

Review of Literature

Hayat et al., (2001) studied the response of hormones in Brassica juncea. The leaves of 30 day old plants of Brassica  juncea  were sprayed with 10−6M aqueous solutions of indole-3-acetic acid (IAA), gibberellic acid (GA), kinetin (KIN), and abscisic acid (ABA) or 10−8 M of 28-homobrassinolide (HBR). All the phytohormones, except abscisic acid, improved the vegetative growth and seed yield at harvest, compared with those sprayed with deionized water (control). 28-homobrassinoslide was most prominent in its effect, generating 32, 30, 36, 70, 25, and 29 % higher values for dry mass, chlorophyll content, carbonic anhydrase activity, and net photosynthetic rate in 60 day old plants, pods per plant, and seed yield at harvest, over the control, respectively. The order of response to various hormones was HBR > GA3 > IAA > KIN > control > ABA.

Hayat et al., (2001) studied the effect of 28-homobrassinolide (HBR) on mustard plant.  Thirty day old plants of mustard (Brassica juncea) were sprayed with 10−10, 10−8, or 10−6 M aqueous solution of 28-homobrassinolide (HBR). The HBR-treated plants were healthier than those treated with water and yielded more. Maximum increase over control was found in 60 day old, 10−8 M-HBR-treated plants in fresh and dry mass per plant, carbonic anhydrase activity, and net photosynthetic rate , at harvest in number of pods per plant and seed yield per plant (the respective values were 25, 30, 34, 69, 24, and 29 %). A further increase in the concentration of 28-homobrassinolid (10−6 M) did not make any additional impact on the growth and yield. Increased carbonic anhydrase activity and photosynthetic rate were correlated with growth and seed yield.

Khan et al., (2002) reported that mustard is cultivated throughout the world for oil in its seeds. It requires high nitrogen input for improved productivity but the nitrogen applied to the soil is not fully utilized by the crops due to various constraints. A field experiment was conducted during 1997–98 in which GA3 (10−5 M) was applied to foliage at 40 day after sowing (pre-flowering) to mustard grown with 0, 40(sub-optimal), 80 (optimal) and 120 (supra-optimal) kg N ha−1. Foliar spray of GA3 was effective only when plants received sufficient N (80 kg N ha−1). GA3 sprays significantly enhanced plant dry mass, leaf area, carbon dioxide exchange rate, plant growth rate, crop growth rate and relative growth rate. GA3 treated plants showed enhanced nitrogen-use efficiency through redistribution of N to seed.

Fariduddin et al., (2003) applied aqueous solutions of salicylic acid (SA) to the foliage of 30 day old plants of mustard (Brassica juncea). The plants sprayed with the lowest used concentration (10−5 M) of SA were healthier than those sprayed with water only or with higher concentrations of SA (10−4 or 10−3 M). 60 day old plants possessed 8.4, 9.8, 9.3, 13.0 and 18.5 % larger dry mass, net photosynthetic rate, carboxylation efficiency, and activities of nitrate reductase and carbonic anhydrase over the control, respectively. Moreover, the number of pods and the seed yield increased by 13.7 and 8.4 % over the control.

Mehra et al., (2003) studied the rate of germination of mustard plant.Seed lots of Brassica juncea was subjected to aerated hydration (AH) for up to 24 h at 20°C. The rate of germination, final germination (% normal seedlings) and germination after the controlled deterioration test increased after AH treatment in seeds that initially had reduced seed quality. The optimum timing of treatment was 12 h .The germination of seed of Brassica juncea was tested before and after 12 h AH at a range of water potentials (0 to -1.00 MPa) and salinities (0-331.8 mmolal NaCl). Both the final normal germination (%) and germination rate, as measured by mean germination time, decreased as water stress (decreased water potential) and salinity increased. However in all cases the normal germination (%) was markedly higher for AH treated seeds. Germination increased at temperatures both above (33-37°C) and below (8-10, 13-14°C) the optimum for germination (22-23°C), thus extending the range of temperatures for successful germination.

Mandal and Sinha (2004) conducted an experiment on sandy loam acidic soil to study the effect of nutrient managements on light interception, photosynthesis, growth, biomass production and yield of Indian mustard (Brassica juncea). Plant height, number of branches per plant, number of siliquae per plant, number of seeds per siliquae, seed and oil yield of Indian mustard improved at 100 % recommended rates of NPK (nitrogen, phosphorus and potassium) .The rate of photosynthesis increased due to appropriate nutrient management treatments with concomitant increase in photosynthetically active radiation, internal CO2 concentration and rate of transpiration and decrease in stomatal resistance. Consequent upon the higher rate of photosynthesis, dry-matter accumulation increased. Thus, significant improvement due to appropriate combination of NPK, borax and ZnSO4 was observed for uptake of nutrients.

Huysen et al., (2004)  reported that mustard (Brassica juncea) plants overexpressing ATP sulfurylase (APS transgenics) were previously shown to have higher shoot selenium (Se) levels and enhanced Se tolerance compared to wild type when supplied with selenate in a hydroponic system. In the present study, these APS (adenosine phosphate sulfyrylase) and  transgenic were evaluated for their capacity to accumulate Se from soil that is naturally rich in Se. Wild-type Indian mustard and the Se hyper accumulator Stanleya pinnata were included for comparison. After 10 weeks on Se soil, the APS transgenic contained 2.5-fold higher shoot Se levels than wild-type Indian mustard, similar to those of Stanleya pinnata. The transgenic contained 40% lower shoot Se levels than wild type. Shoot biomass was comparable for all Indian mustard types and higher than that of Stanleya pinnata. These results obtained with these transgenics on soil are in agreement with those obtained earlier using hydroponics.

Qadir et al., (2004) were studied the effect of cadmium on mustard plant germination. Ten Brassica juncea commonly grown in India were selected for the study to determine their Cd extraction potential and degree of resistance to Cd stress. Ten-day-old seedlings of Brassica juncea were exposed to various levels of cadmium chloride (0.0–2.0 mM) for 72 h in hydroponics culture and leaf samples were analyzed at 24, 48 and 72 h after treatment for the changes in the rate of lipid peroxidation, plant length, biomass accumulation, cadmium accumulation and activities of catalase (CAT), superoxide dismutase (SOD), ascorbate peroxidase (APX) and glutathione reductase (GR) along with ascorbate (Asc) and glutathione contents. A reduction in the plant length, biomass accumulation, CAT activity and ascorbate content was noted in all the cultivars, however, a significant increase in lipid peroxidation rate, Cd accumulation, activities of APX, GR, SOD and glutathione content was observed.

Khan et al., (2005) carried out the research to study the effects of 10~μM GA3 spray on specific leaf area, plant dry mass, leaf carbon dioxide exchange rate (CER), plant growth rate , relative growth rate , net assimilation rate and Sulfur use efficiency of mustard treated with 0, 100 or 200 mg  kg−1 soil levels. Plants treated with 100~mg S kg−1 soil and receiving GA3 treatment showed increased specific leaf area and dry mass accumulation compared to the control. At 0~mg S kg−1soil, N and S concentrations were reduced. They increased with increasing S supply. GA3 application significantly increased N and S concentrations further. A two-fold increase in SUE in GA3-treated plants at 100~mg S kg−1 soil was noted in comparison to the control. SUE was not increased under excess S conditions beyond 100~mg S kg−1 soil. The increase in SUE (sulfur use efficiency) was through increase in the growth, CER (carbon dioxide exchange rate) and use efficiency of Nitrogen by the crop due to GA3 application.

Ashraf and Foolad (2005) were studied the effects of salinity on seed germination. The general purpose of seed priming is to partially hydrate the seed to a point where germination processes are begun but not completed. Treated seeds are usually redried before use, but they would exhibit rapid germination when re‐imbibed under normal or stress conditions. A better understanding of the metabolic events that take place in the seed during priming and subsequent germination will improve the effective application of this technology. The incorporation of advanced molecular biology techniques in seed research is vital to the understanding and integration of multiple metabolic processes that can lead to enhanced seed germination, and consequently to improved stand establishment and crop yield under saline and non‐saline conditions

Belimov et al., (2005) isolated the eleven cadmium-tolerant bacterial strains from the root zone of Indian mustard (Brassica juncea) seedlings grown in Cadmium-supplemented soils as well as sewage sludge and mining waste highly contaminated with Cd. The isolated strains included Variovorax paradoxusRhodococcus and Flavobacterium and were capable of stimulating root elongation of Brassica juncea seedlings either in the presence or absence of toxic Cd concentrations. Some of the strains produced indoles or siderophores, but none possessed C2H2-reduction activity. All the strains, except Flavobacterium strain 5P-3, contained the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase, which hydrolyses ACC (the immediate precursor of plant hormone ethylene) to NH3 and α-ketobutyrate. Variovorax paradoxus utilized ACC as a sole source of N or energy. A positive correlation between the in vitro ACC deaminase activity of the bacteria and their stimulating effect on root elongation suggested that utilization of ACC is an important bacterial trait determining root growth promotion.

Vivek et al., (2005) saw the effect of hexavalent chromium (Cr6+) on Brassica juncea grown for 15 days in hydroponic culture supplemented with 0.2, 2 and 20 μM Cr. The inhibitory response of Cr6+ on growth of Brassica juncea was concentration and time dependent. The stimulation of plant growth, observed in response to exposure to 0.2 μM Cr6+, during initial 5 days was reversed on prolonged treatment and at higher Cr6+concentrations (2 and 20 μM Cr6+). Despite reduction in growth, chlorophyll content increased substantially on 15 days exposure to 20 μM Cr6+. Significant increases in lipid peroxidation and tissue concentration of H2O2 occurred in plants exposed to 2 and 20 μM Cr6+. Effect of Cr6+ on antioxidative enzymes in roots and leaves was differential. The results suggested Cr6+ induced depression in plant growth of Brassica juncea to be a function of increased cellular accumulation of Cr despite increase in the activities of some of the antioxidative enzymes.

Gunasekera et al.,(2006) studied  adaptation of  mustard (Brassica juncea ) to low rainfall, short season, Mediterranean-type environment in south Western Australia by investigating the effects of genotype, environment and their interaction on crop growth and seed yield. Seed yield of mustard and canola in low rainfall environments were higher when sown early in the season (May). Mustards were generally more adapted to stressful environments associated with low rainfall, high temperature and late sowing as inferred from principal component. Mustard produced significantly higher dry matter particularly in later sowings. Average phenotypic stability of mustard genotypes was mainly associated with their greater tolerance to stressful environments (low rainfall, high temperature and late sowing), early vigour, shorter pre-anthesis phases and greater dry matter production.

Danika et al., (2006) a major goal of their selenium (Se) phytoremediation research was to use genetic engineering to develop fast-growing plants with an increased ability to tolerate, accumulate, and volatilize Se. To this end we incorporated a gene (encoding selenocysteine methyltransferase, SMT) from the Se hyperaccumulator, Astragalus bisulcatus, into Indian mustard (Brassica junicea). The resulting transgenic plants successfully enhanced Se phytoremediation in that the plants tolerated and accumulated Se from selenite significantly better than wild type. However, the advantage conferred by the SMT enzyme was much less when Se was supplied as selenate. In order to enhance the phytoremediation of selenate, we developed double transgenic plants that overexpressed the gene encoding ATP sulfurylase (APS) in addition to SMT, i.e., APS × SMT. The results showed that there was a substantial improvement in Se accumulation from selenate (4 to 9 times increase) in transgenic plants over expressing both APS and SMT.

Hayat et al. (2007) studied the change in plant growth, photosynthesis, carbonic anhydrase, nitrate reductase and antioxidative enzymes resulting from the feeding of cadmium and/or 28-homobrassinolide (HBL) to Brassica juncea in 60-day-old plants. One-week old seedlings were supplied with 50, 100 or 150 μM of cadmium along with the nutrient solution. Subsequent seedlings, at day 30, were sprayed with 0.01 μM of HBL to their foliage. The plants fed with cadmium alone exhibited a decline in growth, the levels of carbonic anhydrase and chlorophyll pigments and net photosynthetic rate. Moreover, nitrate content, the activity of nitrate reductase and the level of carbohydrate, both in the leaves and roots, decreased as the concentration of cadmium, in nutrient solution, increased. Compared with the leaves, roots possessed a larger quantity of nitrate. However, the trend was reversed in case of nitrate reductase and the level of carbohydrates in the two plant organs. The toxic effect, generated by cadmium was overcome if the stressed plants were sprayed with HBL.

Bharagava et al., (2008)  studied the metal accumulation potential and its physiological effects in Indian mustard plants (Brassica nigra) grown in soil irrigated with post methanated distillery effluent (25%, 50%, 75%, 100%, v/v) after 30, 60 and 90 days after sowing. An increase in the chlorophyll and protein contents was recorded at the lower concentrations of post methanated distillery effluent (PMDE) at initial exposure periods followed by a decrease at higher concentrations of PMDE compared to their respective controls. An enhanced lipid peroxidation in tested plants was observed, which was evidenced by the increased malondialdehyde content in shoot, leaves and seeds at all the concentrations of PMDE and exposure periods compared to their respective controls. This study revealed that Indian mustard plants (Brassica nigra) are well adopted to tolerate and accumulate high quantities of trace elements due to increased level of antioxidants (cysteine and ascorbic acid) in root, shoot and leaves of the treated plants at all the concentrations and exposure periods except at 90 days, whereas a decrease was observed at 100% PMDE as compared to their respective controls.

Mer et al., (2008) conducted  a green house experiment on mustard plant in the western region of Gujarat State, India to assess its responses to increasing levels of soil salinity mustard was tolerant only to low soil salinity. However, high salt concentrations in the soil reduced the absorption of nitrogen and phosphorus by the young plants. The imbalance of mineral nutrients resulted in a reduction or an inhibition of plant growth.

Parvaiz (2009) studied the effects of salt stress on plant growth parameters, lipid peroxidation and some antioxidant enzyme activities (superoxide dismutase, catalase, peroxidase, and glutathione reductase and ascorbate peroxidase activity) in the leaves of mustard. Plants were exposed to two different concentrations of NaCl stress (100 and 150 mM) for 45 days and were sprayed with GA3 (75 ml pot−1, conc. 75 mg l−1) once a week. Salt stress resulted in decrease in the growth and biomass yield of mustard but the exogenous application of GA3 enhanced these parameters significantly. Application of GA3 counteracted the adverse effects of NaCl salinity on relative water content, electrolyte leakage and chlorophyll (Chl) content. GA3was sufficient to attenuate partially the stimulatory effect of NaCl supply on proline and glycinebetaine biosynthesis. The activity of all the antioxidant enzymes was increased significantly during salt stress in mustard. The exogenous application of GA3 decreased the enzyme activity.

Meetu et al., (2009) showed the toxicity caused by Arsenite (AsIII) and its detoxification responses in two varieties (Varuna and Pusa Bold) of Brassica juncea. Comparisons were made in leaves and roots of both the varieties, which showed that the accumulation pattern in both the varieties were dose and duration dependent, being more in roots for two days and in leaves for four days. Increase/decrease of antioxidant enzymes activities showed not much change at the given concentrations except that the enzyme activities showed significant increase at the lower concentrations. The analysis of isoenzyme pattern in leaves of Pausa Bold showed five and two major bands of SOD (superoxide dismutase) and GPX (glutathio peroxidase) respectively. AsIII treatment leads to the activation of MAP Kinase (mitogen activated protein) activity indicating role of this important cascade in transducing AsIII mediated signals.

Poonum et al., (2009) study was conducted to evaluate the impact of ambient ozone on mustard plants grown under recommended and 1.5 times recommended NPK (nitrogen, phosphorus and potassium) doses at a rural site of India using filtered (FCs) and non-filtered open top chambers (NFCs). Ambient mean O3 concentration varied from 41.65 to 54.2 ppb during the experiment. Plants growing in FCs showed higher photosynthetic rate at both NPK levels, but higher stomatal conductance only at recommended NPK. There were improvements in growth parameters and biomass of plants in FCs as compared to NFCs at both NPK levels with higher increments at 1.5 times recommended. Seed yield and harvest index decreased significantly only at recommended NPK in NFCs. Seed quality in terms of nutrients, protein and oil contents reduced in NFCs at recommended NPK.

Rajpar et al., (2011) studied that humic acid efficiently improves soil fertility and crop productivity, especially on poorly fertile and alkaline-calcareous soils. In this field study, the growth, yield and oil content of three mustard varieties were observed under varying levels of humic acid application to a poorly fertile and alkaline-calcareous soil. The humic acid was applied to soil at the time of sowing in 0, 3.17, 6.35, and 9.35 kg acre-1. Overall varieties, compared to control, the application of humic acid in 6.35 kg acre-1 positively affected almost all the growth and yield parameters. The validity of these results is warranted through further experiments.

Pushp et al., (2013) studied that Soil salinity is one of the most important constraints limiting crop production in arid and semi arid regions. Seed germination is a critical stage in the life history of individual plants and salt tolerance during germination is crucial for the establishment of plants growing in saline soils. The research was carried out in order to test the effect of salinity on germination traits and seedling growth in 25 Indian mustard (Brassica juncea) genotypes. The results revealed statistically significant effects of salinity on germination traits as well as growth characteristics of seedlings. Genotypic responses were significant for germination percentage, speed of germination, germination index, and relative germination rate, which were all generally retarded by salt stress. However, the mean germination time increased under saline conditions. Decline in root/shoot ratio and dry matter of seedlings was observed under salinity.

Akil et al., (2013) studied the effect of microwaves. Over the past few decades with the growth in cellular services all over the world; a noticeable observation comes in front regarding the life span of birds and growth of crops. This is because of obvious reason of increase in microwave presence in our atmosphere. The cellular phones mostly work at 945 MHz frequency. The objective of this study is to investigate the changes in growth rate and germination of mustard plants after exposure of different amount of microwaves in power and duration. The observations for a period of ten days using microwaves treated and untreated soul were carried out. The other control variables such as temperature, humidity, sun light and level of gases (CO2, N2, and O2) were maintained almost same for all the observations. The investigations have shown that the plants grown with microwave exposed soil behaved differently.

Fallah and Baki (2013) to study on effect of NaCl stress on germination and early seedling growth on Brassica juncea. Results indicated that Salinity significantly affected the final germination rate and percent germination and by increasing salinity, significant decrease was observed in percentage of germination and germination rate. As the salinity level increased from 0 to10 M NaCl, more than 80% redaction was observed in seed germination compared to control and it was completely inhibited in higher salinities. According to results, 20°C was the optimal temperature for germination of seeds in salinity situation and any increase or decrease in temperature inhibited germination. Results also showed length of radicle and radicle dry matter significantly was decreased when salinity was increased. Results showed in NaCl concentration at 8 MPa root growth stopped after seed germination and completely inhibited at 10 and 12 MPa.

Methodology

3.0. Site of study:

All the research work is done in the botany research lab of College.

3.1. Plant material:

I have selected the yellow mustard plant (Brassica juncea).  Brassica  juncea is a herbaceous plant with an erect, branched stem up to 1.0 m tall, with a taproot reaching 60-80 cm in depth, lower leaves petioled, green, sometimes with a whitish bloom, ovate to obovate, variously lobed with toothed or frilled edges; upper leaves subentire, short and petioled, constricted at intervals, sessile. The flowers consist of 4 (four) yellow petals arranged in a cruciform manner, 4 (four) yellowish green sepals, a short green pistil with a knobby stigma, and tetradynamous stamens with yellow anthers.  Mustard emerges rapidly (5 to 10 days) and grows rapidly under favorable moisture and temperature conditions. I have taken seeds of three different regions.

3.2. Yellow mustard: Classification

Kingdom: Plantae

Division: Angiospermae

Class: Dicotyledons

Order: Brassicales

Family: Brassicaceae

Genus: Brassica

Species: juncea

3.3. Conditions for germination:

  • 3.3.0. Climate:

Mustard is a cool season crop with short growing season. Adequate water and cool temperatures(less than 85 F) favor a long bloom period.

  • 3.3.1. Soil:

Mustard is adapted best to fertile, well drained, loamy soils, but it can grow in verities of soil type with good drainage. Soils prone to crusting prior to seedling emergence can cause problems. Avoid dry sand, dry, sandy loam soils, also.

  • 3.3.2. Seed germination:

Seed will germinate at soil temperature as low as 40 F.

3.4. Material and method:

  • 3.4.0. Materials:

Mustard seeds of three different regions

Pots

  • 3.4.1. Other materials:

Water

Soil

NaCl

  • 3.4.2. Method:

In order to establish the best priming condition, seeds of the three varieties in study were treated with NaCl solution at 0, 2, 4, 6 and 8 g L-1 for 24 h. After seeds (control seeds) were sown directly in the soil in April 2015. Throughout their vegetative cycles, plants seeds were irrigated with saline water at five levels of NaCl concentrations (0, 2, 4, 6 and 8 g l-1). Germination, root length, and plant height of mustard plant were evaluated with analysis of variance (ANOVA) (p < 0.05) using SPSS software version (17.0). Differences were considered significant at the 5% level (means followed by different letters).

3.5. Germination Record:

Germination was recorded daily. The number of germinated seeds was counted after one week from the start of experiment. Germination percentage was calculated by Close and Wilson (2002) method by using formula:

Germination percentage= number of seeds germinated/total number of seeds grown × 100

3.6. Root length and Plant height:

After 15 days of seed sowing, seedlings from all three varieties were collected. Root length and Plant height was then measured by taking ruler or scale.

Result

4.0. Germination rate:

Germination rate of all the three varieties of mustard plant decreased with increase in salt concentration. Highest germination rate occurred at 0g/l (control) concentration. As the rate of salt concentration increased germination rate decreased.

4.1. Root height:

Significant differences were observed for root length between different varieties of seed. Root length also reduced with the increase in salt concentration. The highest root length was measured at 0g/l control and the lowest root length was measured at 8g/l concentration.

4.2. Plant height:

As concentration of NaCl increased plant height also decreased. Similarly at control concentration highest plant height was measured and minimum at 8g/l. As the salinity level increased from 0 to 8 g/l NaCl, more than 80% redaction was observed in seed germination compared to control and it was completely inhibited in higher salinities.


Table no.1 representing rate of germination of mustard plant

Salt concentration Germination rate Root length Shoot length Plantlet length
gl-1 % cm cm cm
0 88 2 3.8 5.8
2 80 1.8 3.5 5.3
4 70 1.6 3.0 4.6
6 50 1 2.2 3.2
8 12 0.7 1.3 2

 

Table no.2 representing rate of germination of mustard plant

Salt concentration Germination rate Root length Shoot length Plantlet length
g1-1 % cm cm cm
0 92 2.2 3.5 5.7
2 80 1.9 3.2 5.1
4 72 1.4 2.8 4.2
6 55 1 2.4 3.4
8 18 0.5 1.0 1.5

 

Table no.3 representing rate of germination of mustard plant

Salt concentration Germination rate Root length Shoot length Plantlet length
gl-1 % cm cm cm
0 78 2.7 6.7 9.4
2 70 2.4 6.0 8.0
4 62 1.8 5.3 7.1
6 40 1.2 4.1 5.3
8 10 0.8 1.0 1.8

 

Table no.4 mustard plant (after statistical analysis)

ANOVA
Measure
Sum of Squares df Mean Square F Sig.
Between Groups 14902.637 4 3725.659 36.532 .000
Within Groups 1937.668 19 101.983
Total 16840.305 23
Multiple Comparisons
Dependent Variable: measure

LSD

(I) group (J) group Mean Difference (I-J) Std. Error Sig. 95% Confidence Interval
Lower Bound Upper Bound
1 2 -59.60000* 6.77437 .000 -73.7789 -45.4211
3 3.38000 6.77437 .624 -10.7989 17.5589
4 2.24000 6.77437 .745 -11.9389 16.4189
5 .82000 6.77437 .905 -13.3589 14.9989
2 1 59.60000* 6.77437 .000 45.4211 73.7789
3 62.98000* 6.38694 .000 49.6120 76.3480
4 61.84000* 6.38694 .000 48.4720 75.2080
5 60.42000* 6.38694 .000 47.0520 73.7880
3 1 -3.38000 6.77437 .624 -17.5589 10.7989
2 -62.98000* 6.38694 .000 -76.3480 -49.6120
4 -1.14000 6.38694 .860 -14.5080 12.2280
5 -2.56000 6.38694 .693 -15.9280 10.8080
4 1 -2.24000 6.77437 .745 -16.4189 11.9389
2 -61.84000* 6.38694 .000 -75.2080 -48.4720
3 1.14000 6.38694 .860 -12.2280 14.5080
5 -1.42000 6.38694 .826 -14.7880 11.9480
5 1 -.82000 6.77437 .905 -14.9989 13.3589
2 -60.42000* 6.38694 .000 -73.7880 -47.0520
3 2.56000 6.38694 .693 -10.8080 15.9280
4 1.42000 6.38694 .826 -11.9480 14.7880
*. The mean difference is significant at the 0.05 level.

 

Table no 5.

ANOVA
Readings
Sum of Squares df Mean Square F Sig.
Between Groups 14620.954 4 3655.239 21.786 .000
Within Groups 3355.616 20 167.781
Total 17976.570 24
Multiple Comparisons
Dependent Variable: measure

LSD

(I) group (J) group Mean Difference (I-J) Std. Error Sig. 95% Confidence Interval
Lower Bound Upper Bound
1.00 2.00 -59.40000* 8.19221 .000 -76.4887 -42.3113
3.00 2.60000 8.19221 .754 -14.4887 19.6887
4.00 1.42000 8.19221 .864 -15.6687 18.5087
5.00 .02000 8.19221 .998 -17.0687 17.1087
2.00 1.00 59.40000* 8.19221 .000 42.3113 76.4887
3.00 62.00000* 8.19221 .000 44.9113 79.0887
4.00 60.82000* 8.19221 .000 43.7313 77.9087
5.00 59.42000* 8.19221 .000 42.3313 76.5087
3.00 1.00 -2.60000 8.19221 .754 -19.6887 14.4887
2.00 -62.00000* 8.19221 .000 -79.0887 -44.9113
4.00 -1.18000 8.19221 .887 -18.2687 15.9087
5.00 -2.58000 8.19221 .756 -19.6687 14.5087
4.00 1.00 -1.42000 8.19221 .864 -18.5087 15.6687
2.00 -60.82000* 8.19221 .000 -77.9087 -43.7313
3.00 1.18000 8.19221 .887 -15.9087 18.2687
5.00 -1.40000 8.19221 .866 -18.4887 15.6887
5.00 1.00 -.02000 8.19221 .998 -17.1087 17.0687
2.00 -59.42000* 8.19221 .000 -76.5087 -42.3313
3.00 2.58000 8.19221 .756 -14.5087 19.6687
4.00 1.40000 8.19221 .866 -15.6887 18.4887
*. The mean difference is significant at the 0.05 level.

 

Table no.6

ANOVA
readings
Sum of Squares df Mean Square F Sig.
Between Groups 9199.838 4 2299.959 14.815 .000
Within Groups 3104.984 20 155.249
Total 12304.822 24
Multiple Comparisons
Dependent Variable: readings

LSD

(I) groups (J) groups Mean Difference (I-J) Std. Error Sig. 95% Confidence Interval
Lower Bound Upper Bound
1.00 2.00 -48.00000* 7.88034 .000 -64.4381 -31.5619
3.00 2.22000 7.88034 .781 -14.2181 18.6581
4.00 -.62000 7.88034 .938 -17.0581 15.8181
5.00 -2.32000 7.88034 .771 -18.7581 14.1181
2.00 1.00 48.00000* 7.88034 .000 31.5619 64.4381
3.00 50.22000* 7.88034 .000 33.7819 66.6581
4.00 47.38000* 7.88034 .000 30.9419 63.8181
5.00 45.68000* 7.88034 .000 29.2419 62.1181
3.00 1.00 -2.22000 7.88034 .781 -18.6581 14.2181
2.00 -50.22000* 7.88034 .000 -66.6581 -33.7819
4.00 -2.84000 7.88034 .722 -19.2781 13.5981
5.00 -4.54000 7.88034 .571 -20.9781 11.8981
4.00 1.00 .62000 7.88034 .938 -15.8181 17.0581
2.00 -47.38000* 7.88034 .000 -63.8181 -30.9419
3.00 2.84000 7.88034 .722 -13.5981 19.2781
5.00 -1.70000 7.88034 .831 -18.1381 14.7381
5.00 1.00 2.32000 7.88034 .771 -14.1181 18.7581
2.00 -45.68000* 7.88034 .000 -62.1181 -29.2419
3.00 4.54000 7.88034 .571 -11.8981 20.9781
4.00 1.70000 7.88034 .831 -14.7381 18.1381
The mean difference is significant at the 0.05 level.

 

The Effects Of Salinity On The Germination, Root Height, Stem And Planet Height

Fig no.1 representing germination rate of local mustard variety against NaCl concentration.

 Fig no.2 representing germination rate of local mustard variety against NaCl concentration.

Fig no.3 representing germination rate mustard against saline concentration.

Fig no.4 comparison of three varieties

The Effects Of Salinity On The Germination, Root Height, Stem And Planet Height

Fig no.1 of mustard plant

The Effects Of Salinity On The Germination, Root Height, Stem And Planet Height

Fig no.2 of mustard plant

The Effects Of Salinity On The Germination, Root Height, Stem And Planet Height

Fig no.3 mustard

Fig no.4 mustard

Fig no. 5 mustard plant

The Effects Of Salinity On The Germination, Root Height, Stem And Planet Height

Fig no. 6 mustard plant

Discussion

In the present investigation, we focused on evaluation of the potential tolerance of mustard genotypes to salt stress at early stages of growth.As the table no 1,2,and 3represent rate of germination decrease with the increase in salt concentration(0 to 8 g1-1). Seed germination begins with water intake. Salinity prevents water imbibitions, thereby inhibiting the initial process of seed germination (Othman 2005). Salinity imposes osmotic stress by accumulation of Na and Cl ions. Previous studies have shown that increase in salinity delays the initiation of germination leading to reduction in germination percentage. In the present study, seed germination traits were significantly inhibited by salt stress in all the mustard genotypes. Our results are similar to the findings reported by Karagiizel (2003) and later by Li (2008).

Germination index was high under salt stress and comparable with the control. Shanon and Grieve (1999) reported that salinity slowed germination rate at low concentration; the only effect was on germination rate and not on the total percentage of germinating seeds. In further studies with Brassica species (Jamil et al. 2005) and some tomato cultivars (Al-Hirbi et al. 2008) it was demonstrated that germination rates were considerably lower in high salt concentration with respect to the control. Germination time in mustard seeds was considerably affected by salinity.

According to Karagiizel (2003), germination time in different plant species considerably increases with an increase in salt concentration. Salinity influenced germination time more dramatically than the germination percentage (Ozcoban, Demir 2006). This means that increase in salt concentration results in prolongation of germination time. However, salinity caused different germination time delay amongst the cultivars studied. Delayed germination has also been reported by Zapata et al. (2003) on Lactuca sativa and Chartzoulokis and Klapaki (2000) on pepper.

Fooled and Lin (1999) reported that germination under salt stress can be used as a criterion for salt tolerance. Non-significant differences were recorded in mean germination time. Furthermore, the rapid germination may contribute to salt tolerance to some extent.  The seeds of different genotypes of crops may germinate adequately under salt stress. Nevertheless, the seedling may not be fully established for further growth.

Differential inhibition in the root length, shoot length as well as hypocotyl was observed. This phenomenon has been also reported in triticale (Abdul Karim et al. 1992) and wheat (Raiaj-Ahmed et al. 2001; Kaya et al. 2008; El-Hendaway et al. 2011). These authors suggested that seedling parameters are the most important criterion for screening genotypes for salt tolerance at the early growth stages.  Salinity had greater inhibitory effects on seedling growth than on germination, and there was substantial genotypic variation in salt tolerance among the mustard genotypes. The significant reduction in seedling growth by salinity may be attributed to the toxic effect of sodium chloride and unbalanced nutrient uptake by seedlings. These deleterious effects of salinity may result in a significant decrease in photosynthesis and increase in respiration rate leading to a shortage of assimilate to the developing organs, thus slowing down growth or stopping it entirely (El-Hendaway et al. 2005).

As salinity enhances osmotic pressure leading to reduction in water absorbance, cell division and differentiation are inhibited, which adversely affects metabolic and physiological processes. This causes more delay in initiation of germination followed by prolonged seed germination duration (Kang, Saltvett 2002) and ultimately reducing plumule and radical length (Keshavarzi 2012). Similar results have been reported by Etesami and Galeshi (2008) for barley, Massai et al. (2004) for Prunus and El-Hendawy et al. (2011) for wheat.

Salinity significantly (p <0.05) affected the final germination rate and percent germination of Brassica juncea as table no.4, 5 and 6 indicated. These results are similar in line with Jeannette et al. (2002) and Okcu et al.(2005). They found that the mean time to germination of Phaseolus and Brassica species increased with the addition of NaCl and this increasing was greater in higher concentration as compared to low concentration. The highest percentage of seed germination was recorded in distilled water and germination percentages decreased sharply with increasing NaCl concentration. It is also assumed that in addition to toxic effects of certain ions, higher concentration of salt reduces the water potential in the medium which hinders water absorption by germinating seeds and thus reduces germination (Jamil et al. 2006).

It is indicated that germination rate and the final seed germination decrease with the decrease of the water movement into the seeds during imbibitions (Jamil et al. 2005). Salinity stress can affect seed germination through osmotic effects. Salt induced inhibition of seed germination could be attributed to osmotic stress or to specific ion toxicity (Huang and Redmann 1995).

Effects of NaCl stress on seed germination of 98 genotypes of Brassica juncea were investigated by Kuhad et al. 1989. NaCl stress types significantly reduced germination percentage, dry matter weight, shoot and root length of seedlings. In Indian mustard (Brassica juncea) increasing salinity levels of irrigation water progressively reduced the seed germination of six cultivars (Ray and Khaddar 1990).

Seeds have the highest resistance to extreme environmental stresses, whereas germination is considered as the most sensitive stage and seedlings are most susceptible in the life cycle of a plant (Sidari et al. 2008).

Therefore, successful establishment of a plant population is dependent on the adaptive aspects of seed germination and of early seedling growth (Qu et al. 2007).

Concentrations of 8-10 MPa NaCl significantly decreased hypocotyls elongation (P≤ 0.05), whereas concentrations of 12 MPa completely inhibited them (P≤ 0.05). No hypocotyl elongation occurred at NaCl concentrations of ≥10 MPa.

Parti et al. (2003) reported in Brassica juncea higher levels of salinity adversely affected plant growth, seed yield and total lipids of seeds.Radicel elongation also significantly affected by NaCl osmotic stress. Results showed in NaCl concentration at 8 MPa root growth stopped after seed germination and completely inhibited at 10 and 12 MPa. Reduction in root growth under saline conditions may either be due to decrease in the availability of water or increase in sodium chloride toxicity.

Conclusion:

As the result indicate that as the salt concentration increased from 0g/l to 8g/l then rate of germination decreased as well as root height, stem height and plant height decreased. Plant show maximum growth at control concentration (0g/l) and minimum growth at 8/l.

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