response of seed priming with boron on ... of...broccoli seeds cv. marathon of sakata seed company...

International Journal of Agricultural Science

and Research (IJASR)

ISSN 2250-0057

Vol. 3 Issue 2, Jun 2013, 183-194

© TJPRC Pvt. Ltd.

ISSN 2249–6939

Vol.2, Issue 2 (2012) 1-16

© TJPRC Pvt. Ltd.,

RESPONSE OF SEED PRIMING WITH BORON ON GERMINATION AND SEEDLING

SPROUTS OF BROCCOLI

NOOR-UN-NISA MEMON1, MOULA BUX GANDAHI

1, VAJANTI MALA PAHOJA

2 & NASIM SHARIF

3

1Department of Horticulture, Sindh Agriculture University, Tandojam, Sindh, Pakistan

2Department of Crop Physiology, Sindh Agriculture University, Tandojam, Sindh, Pakistan

3Ayub Agriculture Research Institute, Faisalabad, Punjab, Pakistan

ABSTRACT

Better germination and vigor of the seedlings are main problems for the success of stand establishment of crop

plants. The present study was therefore conducted on seed priming with boron to observe the efficacy of priming on

germination and growth related attributes of the broccoli seedlings. Broccoli seeds (cultivar Marathon) of SAKATA Seed

Company were soaked in boric acid solution at 0.01, 0.05, 0.5 and 1% (w/v) for 18 hours. Seeds were also soaked in

distilled water (hydropriming) and unprimed seeds were taken as control. All primed and non-primed seeds were grown in

earthen pots contained sandy clay loam soil with EC 0.74 dS/m, pH 7.8, organic matter (0.71%), total nitrogen content

(0.06%), available phosphorus 3.6 mg kg-1

, exchangeable potassium 168 mg kg-1

and Boron 0.48 ppm. The results revealed

that Germination percentage (GP), Mean germination time (MGT), Germination index (GI), Seedling vigor index (SVI),

Chlorophyll content, Shoot and root related attributes were significantly influenced by primed seeds as compared to

unprimed seeds. The highest germination index (6.289), seedling vigor index (1753.3), chlorophyll content (4.137 mg ml-1

)

and less mean germination time (3.23 days), maximum length of shoot (5.97 cm), root (11.57 cm), weight of the shoot

(15.35 g) and root (2.68 g) were observed from the treatment where seeds were primed with boron solution at the lowest

concentration of 0.01%.

KEYWORDS: Seed Priming, Boron, Germination, Seedling Vigor

INTRODUCTION

Broccoli (Brassica oleracea) is one of the main cole crops of Brassicaceae or cruciferae family. It is supposed to

be the first of the Cole crops evolved from the wild species of cabbage or kale (Rubatzky and Yamaguchi, 1997). Broccoli,

a rich source of vitamin C, E, B1, carotenoids, phenolic and possess anticancer properties (Goncalves et al., 2011).

Consumption of broccoli is increasing day by day as staple food due to its health promoting properties. Broccoli is green in

color due to chlorophyll within the sepals of the floral buds and grown as an annual crop for its immature flower buds

(Gray, 1982; Jett et al., 1995). In most of the annual crops yield losses occur due to poor germination rate and stand

establishment of the crop (Eskandari and Kazemi, 2012). Uniform and better stand establishment of the crop may allow

grower to reduce costs and increases harvest efficiency (Heather and Sieczka, 1991).

Better germination and vigor seedlings are the main basic foundations for the success of stand establishment of

any crop plant. Various seed technologies including priming, coating and conditioning of the seeds are applied to improve

germination and emergence in seeds of many crops (Arif et al., 2008; Taylor et. al., 1998). Seed priming is a technique in

which seeds are soaked in solutions of low water potential that initiates pre-germinative metabolic activity but prevents

radical protrusion (Bradford, 1986; McDonald, 2000; Ashraf and Foolad, 2005; Farooq et al., 2006; Janmohammadi et al.,

2008). However, the beneficial effects of seed priming have been well documented in many research based published data

184 Noor-Un-Nisa Memon, Moula Bux Gandahi, Vajanti Mala Pahoja & Nasim Sharif

include early germination; improves germination rate (Dahal et al., 1990; Jett et al., 1996; Bradford, 1986; Taylor and

Harman, 1990; Ghassemi-Golezani et al., 2008); breaks dormancy (Cantliffe et al., 1984; Wurr and Fellows, 1984)

improves seedling vigor (Harris, 1996; Saber et al., 2012); strengthens stand establishment (Khan et al., 1992; Jett et al.,

1995; Arif et al., 2005; Ali et al., 2007; Diniz et al., 2009) and increase yield (Rengel and Graham, 1995a, 1995b; Yilmaz

et al., 1998). Besides all these beneficial effects, seed priming also reduce leakage of metabolites, repairs deteriorated seed

parts, improve RNA and protein synthesis (McDonald, 2000).

Number of priming treatments are generally in use including hydro, halo, osmo, thermo and matrix priming

(Khan, 1992; McDonald, 2000; Basra et al., 2005; Ghiyasi et al., 2008). Every technique responds differently in various

crops. These various priming technique have been successfully applied for their beneficial effects in many agronomic

crops including wheat, sugar beet, maize, soybean and sunflower, rice (Khajeh-Hosseini et al., 2003; Sadeghian and

Yavari, 2004; Iqbal et al., 2012; Rehman et al., 2012; Ahmad et al., 2012). However very few studies have also been

conducted in vegetable crops including summer squash (Atta Aly, 1998); common beans (Mohondas, 1985); water melon

(Elmstrom, 1985); carrot and onions (Brocklehurst and Dearman, 1983); okra (Mereddy et al., 2000; Shah et al., 2011);

tomato (Afzal et al., 2011; Nawaz et al., 2011); hot pepper (Amjad et al., 2007); lentil (Saglam et al., 2010) and Dil

(Mirshekari, 2012).

Nutripriming is now recently focused by using macro or micronutrient enriched seeds as reported by Rehman et

al. (2012) and Mirshekari (2012). Generally nutrients are delivered to plants as soil applications, fertigation or foliar spray

(Johnson et al., 2005; Rober, 2008). Applying nutrients as seed treatment, through seed coating and seed priming, is

another option which avoids aforementioned risks (Farooq et al., 2012). The use of micronutrient-enriched seeds (seed

priming) has been reported to be a better strategy in overcoming micronutrient deficiencies (Musakhandov, 1984; Harris et

al. 1999). Priming the seeds with micronutrients makes them able to rapidly imbibe water and revive metabolism and

germination. This then results in a higher germination rate (Rowse, 1995), improved stand establishment, increased

drought and pest tolerance, and ultimately higher yields (Harris et al., 1999).

Boron is one of the essential micronutrient for all vascular plants (Warington, 1923). It is absorbed from the soil

by plants as borate. Since boron is non-mobile in plants, and continuous supply from soil or planting media is required in

all plant meristems. Deficiency or toxicity of boron causes severe reduction in crop yield, which is due to disturbance in

metabolic events involving boron (Brown et al., 2002). Boron has vital role in the formation of proteins, nitrogen

metablolism, cell division, cell membrane integrity, cell wall formation, nucleic acids, and antioxidative systems (Parr and

Loughman, 1983; Gupta et al., 1985; Cakmak and Ro¨mheld, 1997; Goldbach and Wimmer, 2007; Bonilla et al., 2009;

Koshiba et al., 2009). It has also vital role in the process of transpiration for the movement of potassium to the stomata of

the leaf. Boron also maintains stable balance between sugars and starches, pollination and seed production (Gupta et al.,

1985). Seed priming with macro or micro nutrient application is recently done on various agronomic crops including dill

(Mirshekari, 2012), rice (Kalita et al., 2002; Johnson et al., 2005), maize (Harris et al., 2007); wheat and lentil (Jhonson et

al., 2005, Iqbal et al., 2012). There is no report reported in vegetables except dill (Mirshekari, 2012).

MATERIAL AND METHODS

The experiment was carried out at horticulture garden, Department of Horticulture Sindh Agriculture University,

Tandojam, Pakistan located at 25o 25‟60‟N 68

o31‟ 60E, altitude 19.5 m asl. Broccoli seeds CV. Marathon of SAKATA

seed company were used in the present study to observe the efficacy of seed priming with boron on germination and

growth attributes of seedlings. Twenty broccoli seeds were soaked in solution of 0.01, 0.05, 0.5 and 1% boric acid solution

Response of Seed Priming with Boron on Germination and Seedling Sprouts of Broccoli 185

(w/v) for 18 hours. Seeds were also soaked in distilled water (hydropriming) and unprimed seeds were taken as control.

The seeds were surface washed thrice with distilled water and dried at room temperature. All primed and unprimed seeds

were planted in earthen pots containing soil. The soil was analyzed for soil texture by Bouyoucos Hydrometer method, pH

of 1:5 soil-water extract by pH meter, EC of 1:5 soil-water extract by EC meter, organic matter by Walkley-Black, total

nitrogen by Kjeldahl's method, available phosphorus by Olsen, exchangeable potassium by extraction with ammonium

acetate and boron by hot water method. The soil was sandy clay loam, non-saline in nature with EC 0.74 dS/m and slightly

alkaline in reaction with pH 7.8. The soil was also low in organic matter (0.71%), total nitrogen content (0.06%), available

phosphorus 3.6 mg kg-1

, exchangeable potassium 168 mg kg-1

and Boron 0.48 ppm. Monthly mean minimum and maximum

temperatures and other related data of weather was also recorded and presented in “Table 1”.

The data was recorded for Germination Percentage (GP), Mean Germination Time (MGT), Germination Index (GI),

Seedling Vigor Index (SVI), length and fresh weight of shoot and root, electrolyte leakage of leaf and chlorophyll content of

broccoli sprouts.

Germination percentage -Germination was checked on every alternative day for six days and the germination

percentage was calculated by using following equation as described by Larsen and Andreasen (2004).

GP = Σn/ N ×100

where n is number of germinated seeds at each counting and N is total seeds in each treatment.

Mean Germination Time (MGT) was calculated by using following equation of Ellis and Roberts (1981)

MGT = ∑ Dn

∑n

where n is the number of seeds germinated on day D and D is the number of days as counted from the beginning of

germination.

Germination index (GI) was calculated by the formula given by the Association of Offi cial Seed Analysts (1983)

GI = Number of germinated seeds + ---------- + ---------- + Number of germinated seeds

Days of first count Days of last count

Seedling Vigor Index (SVI) was calculated by using following formula, Abdul-Baki and Anderson (1970)

Vigor index (VI) = [seedling length (cm)× germination percentage]

One and half months old seedlings were kept under observation for length and weight of shoot and root, electrolyte

leakage of leaf and chlorophyll content of leaf.

Electrolyte leakage of leaf was measured by taking leaf discs of size 1 cm2 and weight 0.5 g from random

samples of leaf. The leaf discs were rinsed well in deionized water prior to incubation in 25 ml of deionized water for 3 h at

room temperature. After incubation, the conductivity (value A) of the bathing solution was measured with the conductivity

meter. The petal discs were boiled with bathing solution for 15 min to lyse all cells. After cooling to room temperature, the

conductivity (value B) of the bathing solution was again measured. The electrolyte leakage was expressed as percent value

according to the formula given below.

Electrolyte leakage of leaf % = (Value A/Value B) 100

Chlorophyll content – leaf sample of 0.5 g was grinded in 10 mL of 80% acetone and filtered with Whatman paper 1


by adding more 10 ml of acetone. Samples were poured in cuvettes at 663 and 645 nm using UV-Vis spectrophotometer

(Bruinsma, 1963)

The experiment was carried out in Completely Randomized Design with three replications. The obtained data was

statistically analyzed by using software Statistixs 8.1 and interpreted on the basis of Duncan‟s Multiple Range Test

(DMRT).

Table 1: Monthly Mean Temperatures and Other Related Weather Data Recorded During the Study

Months/Year Rainfall (mm) Temperatures (0

0C) Relative Humidity

(%)

Cloudness

(Octas)

Sunshine

(Hours) Minimum Maximum

December, 2012 0 10.6 25.7 55 0.3 7.9

January, 2013 0.03 10.5 25 54 0.7 8.1

Source: Regional Agro Met Centre Tandojam, Sindh, Pakistan

RESULTS

Germination percentage was significantly influenced by seed priming with boron in relation to number of days

“Figure 1”. Unprimed seeds took more time for germination as compared to hydro and boron primed seeds producing 75%

germination on 6th

day of plantation with no germination on 2nd

day of plantation. However, no germination was also

observed from primed seeds on 2nd

day of plantation but most part of the germination was covered on 4th

day of plantation

producing the highest germination (98.33%) from seeds primed with boron solution at concentration of 0.01% as compared

to unprimed seeds (13.33%). On the same 4th

day of plantation, germination was observed decreased with increasing

concentrations of boron i.e. from 0.05 to 1.0%. However 100% germination observed form the seeds primed with boron at

concentration of 0.01 and 0.05% each on 6th

day of plantation. These results were at par with the results obtained from

hydroprimed seeds.

On the basis of mean germination as presented in Figure 2, the highest mean germination (66.11%) was observed

from the seeds primed with boron at concentration of 0.01 “Figure 2”. These results were at par with the results obtained

from hydroprimed seeds (61.11%) and primed seeds with boron at concentration of 0.05% (61.67%). The lowest mean

germination (25%) was recorded from unprimed seeds.

Germination index (GI), mean germination time (MGT), seedling vigor index (SVI) and chlorophyll content was

highly influenced by boron seed priming “Table 2”. The highest germination index (6.289), seedling vigor index (1753.3),

chlorophyll content (4.137 mg ml-1

) and less mean germination time (3.23 days) were observed from the treatment where

seeds were primed with boron solution with the lowest concentration of boron i.e. 0.01%. The results of all these traits are

statistically at par with the results obtained from the treatment where seeds were primed with boron solution at

concentration of 0.05%. Moreover, results were observed decreased with increasing concentration of boron solution i.e.

0.5 to 1.0%. Better results were also observed from hydroprimed seeds as compared to unprimed seeds. No effect of

priming was observed on electrolyte leakage of leaf.

Shoot and root related parameters were significantly influenced by seed priming with boron “Table 3”. The

maximum length of shoot (5.97 cm), root (11.57 cm), weight of the shoot (15.35 g) and root (2.68 g) was observed from

the treatment where seeds were primed with the lowest concentration solution of boron (0.01%). However only weight of

the shoot and root had non-significant results with the results produced from the seeds primed with boron solution at 0.05%

“Table 3”.


HP= Hydroprimed Seeds; BP= Boron Primed Seeds

Figure 1: Effect of Priming on Germination Percentage of Broccoli Seeds in Relation to Number of Days

HP= Hydroprimed Seeds; BP= Boron Primed Seeds

Figure 2: Effect of Priming on Mean Germination of Broccoli Seeds

Table 2: Influence of Seed Priming with Boron on Germination Related Parameters, Chlorophyll and

Electrolyte Leakage of Leaf Content

Priming Treatments

(Hydro and Boron Primed) GI MGT (Days) SVI

Chlorophyll Content

(mg ml-1

)

Electrolyte

Leakage

of Leaf

(%)

Unprimed seeds (control) 2.989 e 5.097 a 753.67 e 2.999 bc 11.83

Seeds primed with Distilled water 5.761 bc 3.65 bc 1260.0 bc 3.605 ab 10.82

Seeds primed with B @ 0.01% 6.289 a 3.23 c 1753.3 a 4.137 a 14.51

Seeds primed with B @ 0.05% 6.017 ab 3.45 c 1413.3 b 3.867 a 11.28

Seeds primed with B @ 0.5% 5.450 c 3.62 bc 1124.0 c 2.885 c 13.1

Seeds primed with B @ 1.0% 4.555 d 4.03 b 946.50 d 3.018 bc 12.57

MGT= Mean Germination Time, GI = Germination Index, SVI = Seedling Vigor Index

Table 3: Response of Seed Priming with Boron on Length and Weight of the Shoot and Root of the

Seedlings

Treatments Shoot Length (cm) Root Length (cm) Shoot Fresh

Weight (g)

Root Fresh

Weight (g)

Unprimed seeds (control) 3.67 b 6.17 e 7.92 bc 1.31 b

Seeds primed with Distilled water 3.97 b 8.63 c 10.46 b 1.50 b

Seeds primed with B @ 0.01% 5.97 a 11.57 a 14.11 a 2.68 a

Seeds primed with B @ 0.05% 4.37 b 9.77 b 15.35 a 2.33 a

Seeds primed with B @ 0.5% 3.83 b 7.77 cd 7.30 bc 1.65 b

Seeds primed with B @1.0% 3.90 b 7.50 d 6.52 c 1.42 b


DISCUSSIONS

Seed priming with boron (nutripriming) exhibited beneficial effects for seed germination, mean germination time,

germination index, seedling vigor index chlorophyll content of leaf, and shoot root related parameters. Priming enhanced

seed performances are related to the repair and the buildup of nucleic acid, enhanced synthesis of protein, repair of

membranes and improves antioxidant system (McDonald, 1999; Hsu et al., 2003). The beneficial effects of priming with

nutrients have been successfully reported by various scientists in various crops (Peeran and Natanasabapathy, 1980,

Sherrell, 1984, Wilhelm et al., 1980; Shah et al., 2011; Aboutalebian et al., 2012; Mirshekari et al., 2012; Rehman et al.,

2012). However, deficiency or toxicity of these nutrients may damage seed or restrict germination as reported by Roberts

(1948). It may also cause abnormal seedlings as reported by Louzada and Vieira (2005) in bean seeds. They observed

mortality of the seedlings in bean seeds due to high applications of micronutrients. Diniz et al. (2009) observed reduced

percentage of germination when higher doses of micronutreients were used for priming in sweet pepper. The low

concentration of boron has been reported to activate key enzymes including phosphorylase, a-amylase etc involved in

starch metabolism. High concentration of boron may be toxic as reported by Bonilla et al. (2004).

In present study, germination was observed decreased with increased doses of boron i.e from 0.5 to 1%. Rehman

et al. (2012) reported no germination from rice seeds primed with boron at 0.5%. While they observed better results for

stand establishment of the crop from the seeds primed with boron at 0.001 and 0.1% concentration. Ajouri et al. (2004)

observed significantly reduced germination in barley when they applied boric acid exceeding 0.04 M. However in papaya,

substantial improvement in germination and early seedling growth was observed when seeds were primed in 2 mg L-1

B

solution for 6 h (Deb et al., 2010). The higher doses of micronutrients in soil solution may also slow the establishment of

the seedlings as reported by Mershikari (2012).

Seedling vigor index is important for rapid stand establishment and early growth of the plants as reported by

Tabrizian and Osareh (2007). Mirshekari (2012) observed restricted seedling vigor index of dill when he used boron and

iron concentrations beyond 1% and 1.5% respectively. He observed higher SVI at 1% boron solution. However, further

SVI increased when boron was used in combination with iron. In present study, SVI was observed higher at the lowest

concentration of boron.The significant increase in shoot and root lengths and fresh weight from seeds primed at the lowest

concentration of boron may be due to its involvement in cell elongation or cell division and meristematic growth

(Bohnsack and Albert 1977; Shelp, 1993; Mouhtaridou et al. 2004; Khan et al. 2006). Boron at lowest concentration of

0.001% has been found successful for improved lengths and fresh weights of shoot and root in rice cultivar as reported by

Rehman et al. (2012). Increasing levels of boron may decrease the fresh weight of the shoot as reported by Ayvaz et al.

(2012). In contrast, Kumar et al. (2008) reported increased plant height of pea when seeds were primed with boron at 0.5%.

This variation in results may be of different genotypes, boron application method, and other soil and environment related

conditions. The results of the present study regarding chlorophyll content is the confirmation that boron is necessary for

increased chlorophyll content. These results are in agreement with the results of Rehman et al. (2012) and Mouhtaridour et

al. (2004). Rehman et al. (2012) reported increased chlorophyll content of leaf from priming solution had very low levels

of boron (0.001%). However, deficiency of boron caused reduction in chlorophyll content as reported by Kastori et al.

(1995). Deficiency or toxicity of boron levels are very narrow in plants (Çelik et al.,1998) and may vary from crop to crop.

CONCLUSIONS

It is concluded from the present study that seed priming with boron is beneficial to improve germination and other

growth related attributes of the seedlings. However the increased levels of boron caused reduction in most of the evaluated


parameters of broccoli seedlings. Further experimentation is required to optimize boron concentrations for each specific

cop.

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