Seed Hardening, Foliar Spraying and Their Combined Effect on Dry Matter Partitioning in Green Gram (Vigna radiata L.)

Mung bean (Vigna radiata L.) is a drought resistant crop and suitable for dry land farming and predominantly used as an intercrop with other crops. It is a very good catch crop in summer and can be grown very well in this season. Mung bean is a short duration, low input requiring crop that matures in 65 to 80 days, photo and thermo-insensitive in nature. However, the productivity of mung bean is low.
 
Efforts made to maximize yield, is largely hampered by adverse effect of abiotic stress such as salinity and drought. These effects cause a huge loss due to low yield and failure of the crop to establish in some cases. Pre-sowing hardening seed treatment is an easy, low cost and low risk technique and also an alternative approach recently used to overcome the effect of abiotic stresses in agricultural production. It is found to be efficient in improving seed emergence and growth of crops (Sankar Ganesh et al., 2013). It was reported clearly that the hardening treatment enhance seeds vigour by protecting structure of the plasma membrane against injury during stress (Bewley and Black, 1982; JunMin et al., 2000). It is a well established fact that, pre-soaking seeds with optimal concentration of phytohormones enhance their germination, dry matter accumulation and partitioning and yield of some crop species under condition of environmental stress by increasing nutrient reserves through increased physiological activities and root proliferation (Bozeuk, 1981).
 
Considering the constraints in the production potential of mung bean it is worthwhile to study the influence of different seed hardening and foliar spraying treatments on the production potential of mung bean. It is also of utmost importance to understand the physiological basis of dry matter accumulation, its partitioning in various plant parts and thereby yield variation due to seed hardening and foliar spraying of various growth regulators and chemicals. The pre-requisite for higher yield is related with the ability of genotype to produce high amount of total dry matter. The manner in which the net dry matter is produced and distributed among the different parts of plant will determine the economic yield (Patil et al., 2007). The present study was therefore, undertaken to assess the pattern of dry matter accumulation and its partitioning in various plant parts in relation to yield in green gram.
 

The present work was carried out at Agronomy farm, Anand Agricultural University, Anand to study the effect of seed hardening, foliar spraying and their combined effect on dry matter accumulation and its distribution in various plant parts in green gram (Vigna radiata L.) during summer season of 2015-16 and 2016-17. The trial was laid out in a randomized block design with three replications and sixteen treatment combinations including five seed hardening treatments, five foliar spraying treatments, five seed hardening treatments with foliar spraying and one absolute control treatment. Seeds of mung bean var. GAM- 5 were imposed with the following seed treatments.
 
The different solutions with different concentrations viz., CaCl₂ 2%, Cycocel 500 mg l^-1 & 1000 mg l^-1 and  NAA 25 mg l^-1 & 50 mg l^-1 were used in this experiment for seed hardening. The solutions were prepared by dissolving 10 g of CaCl₂ directly in the water while 250 mg & 500 mg of Cycocel and 12.50 mg & 25.00 mg of NAA in small amount of ethyl alcohol separately and after dissolved completely, made the final volume up to 500 ml by addition of water.
 
Seed hardening treatments were given to sufficient quantity of seeds of Mung bean cv. GAM-5. For hardening, seeds were soaked in above prepared various solutions of double the volume of seed for three hours. This will ensure that seeds remained immersed in the solution, so as to avoid precocious germination during the treatment period. Hardening was given in flasks under room temperature. The seeds were then removed from respective solutions and kept overnight in shade for drying to attain the seeds to its original moisture level. The seeds were ready for sowing in field on next day.
 
The different solutions with their different concentrations were used in this experiment for foliar spraying. Stock solutions of  CaCl₂ 1%, Cycocel 500 mg l^-1 & 1000 mg l^-1  and  NAA 25 mg l^-1 & 50 mg l^-1 were prepared by dissolving 100 g of CaCl₂ directly in the water while 5.0 g & 10.0 g of Cycocel and 250 mg & 500 mg NAA  in small amount of ethyl alcohol separately and made the total volume up to 500 ml or 1 litter by addition of water as stock solution for each agro chemical and PGRs for better handling from lab to field level for spraying. Working solution was prepared by diluting each stock solution with water and made final quantity of 10 litter of each solution. The spraying was carried out as per treatments during the morning time or before noon at 30 days after sowing (DAS) in respective gross plot of each replication using knapsack sprayer.
 
 
Treat. no.            Treatment details
 
      T₁        CaCl₂ 2% seed hardening (SH)
      T₂        CCC 500 mg/L seed hardening (SH)
      T₃        CCC 1000 mg/L seed hardeningg (SH)
      T₄        NAA 25 mg/L seed hardening (SH)
      T₅        NAA 50 mg/L seed hardening (SH)
      T₆        CaCl₂ 1% spraying at 30 days after sowing (DAS)
      T₇        CCC 500 mg/L spraying at 30 days after sowing (DAS)
      T₈        CCC 1000 mg/L spraying at 30 days after sowing (DAS)
      T₉        NAA 25 mg/L spraying at 30 days after sowing (DAS)
      T₁₀       NAA 50 mg/L spraying at 30 days after sowing (DAS)
      T₁₁       CaCl₂ 2%  seed hardening (SH) + 1% spraying at 30 days after sowing (DAS)
      T₁₂       CCC 500 mg/L seed hardening (SH) + spraying at 30 days after sowing (DAS)
      T₁₃       CCC 1000 mg/L seed hardening (SH) + spraying at 30 days after sowing (DAS)
      T₁₄       NAA 25 mg/L seed hardening (SH) + spraying at 30 days after sowing (DAS)
      T₁₅       NAA 50 mg/L seed hardening (SH) + spraying at 30 days after sowing (DAS)
      T₁₆       Absolute control
 
Crop management
 
Right after sowing, the experimental plots were irrigated regularly at every 10 to 15 days interval until pods matured. Weeding was done manually. Plant protection measures applied as and when required.
 
Dry matter accumulation
 
Sampling was done at every 15 days interval from 30 DAS until harvest regardless of growth stage. The total dry matter production was recorded by destructive method. Randomly selected five plant samples were uprooted and separated into root, leaf, stem and reproductive parts and dried in oven at 80°C until constant weight was obtained. Total dry matter was calculated by adding dry weight of different plant parts and expressed as grams per plant at different intervals 30, 45, 60 and at harvest of crop growth period.

Dynamics of dry matter accumulation
 
The dry matter production is the product of net accumulation of photosynthesis. The accumulation of photosynthesis in various plant parts depends upon the net assimilation rate, the duration of sunshine and the photosynthesizing efficiency of genotype. The manner in which the dry matter is produced by the plant and distributed among its different parts is important for recording good yield (Patil et al., 2007).
       
Both treatments and sampling time i.e. growth stage had a significant effect on dry matter accumulation (g plant^-1) (P < 0.05). Production of dry matter was slow during the first month after sowing, which is typical for legumes adapted to winter conditions (Zapata et al., 2019). The data on dry matter at different intervals (Table 1 to 5) revealed that the dry matter accumulation in leaves, stem, root and pods showed independent behaviour over the crop growth period. In early growth phase larger portion of total dry matter was shared by the leaves than the stem. The dry matter accumulation in pods initiated after 30 DAS with reproductive parts like flowers and continued to increase upto the harvest.










       
In the present study, significant differences were observed in the dry matter per plant due to seed hardening, foliar application and their combined effect treatments (Table 1 to 5).
 
Leaf dry matter accumulation at various growth stages
 
In the present investigation, leaf dry matter per plant indicated significant differences due to seed hardening and foliar spraying treatments over absolute control. When the treatments were compared, seed hardening with 2% CaCl₂ (T₁₁ and T₁) recorded significantly higher leaf dry weight per plant (2.060, 2.470, 2.265 g and 2.053, 2.453, 2.253 g) while minimum leaf dry matter per plant was observed (1.630, 1.940 and 1.785 g) with untreated absolute control (T₁₆) at 30 DAS during summer 2016, 2017 seasons and on pooled basis respectively. At 45 days after sowing, significant differences were noticed for the leaf dry matter per plant. The treatment with CaCl₂ 2% seed hardening + 1% spraying at 30 DAS (T₁₁) noted the highest leaf dry matter accumulation per plant at 45 DAS (5.000, 5.343 and 5.172 g), (7.027, 7.620 and 7.323 g) at 60 DAS  and (4.023, 4.357 and 4.190 g) at harvest during 2016, 2017 and in pooled analysis, respectively (Table-1). Seed hardening and foliar spraying with CaCl₂ increased leaf dry weight. This might be due to metabolic changes like high level of synthetic reaction even during drought; leaves of hardened plants have more starch, higher rate of photosynthesis because of increase in the bound water and higher organic phosphorous and nucleoproteins. These results are conformity with findings of Misra and Dwivedi (1980) and Gurudev Singh et al., (1991) in wheat, Kinjal (2017) in black gram and Corleto et al., (1977) in green gram. The leaf dry matter decreased in all the treatments after 60 DAS at the time of pod development upto harvest. The dry matter accumulation in leaf increased from 30 to 60 DAS and declined thereafter due to shading and leaf senescence.
Stem dry matter accumulation at various growth stages
 
It was found that seed hardening and foliar application of agro chemical CaCl₂, growth retardant (cycocel) as well as growth promoter (NAA) increased stem dry matter per plant as compared to absolute control treatment. The treatment with CaCl₂ 2% seed hardening + 1% spraying at 30 DAS (T₁₁) recorded highest stem dry matter accumulation per plant (4.567, 5.027 and 4.797 g) at 45 DAS, at 60 DAS (7.600, 8.050 and 7.825 g) and (8.350, 8.813 and 8.582 g) at harvest during 2016, 2017 and in pooled analysis, respectively. The absolute control treatment (T₁₆) registered lowest stem dry matter partitioning during all growth stages (Table 2). The treatment of calcium chloride, NAA leads to redistribution of nutrient reserves which results in the greater internodal length, while Cycocel reduced intermodal length but increased number of branches and thereby increases the stem dry weight. Increased in stem dry weight by seed hardening was also reported by Misra and Dwivedi (1980) and Gurudev Singh et al., (1991) in wheat, Kinjal (2017) in black gram and Corleto et al., (1977) in green gram. There was a very less rate of increase in the stem dry matter from pod development stage after 60 DAS till the harvest leading to an increase in pod weight.
 
Root Dry Matter Accumulation at various growth stages
 
Among the treatments, seed hardening with NAA 50 mg/L (T₁₅ and T₅) recorded numerically more root dry weight per plant (0.485, 0.535 and 0.482, 0.533 g) at 30 DAS in summer 2016 and 2017 seasons, respectively. But on pooled basis NAA 50 mg/L (T₁₅ and T₅) recorded significantly higher root dry weight per plant (0.510 and 0.507 g) at 30 DAS. The lowest root dry weight per plant was recorded (0.400, 0.463 and 0.432 g) during 2016, 2017 and in pooled, respectively with absolute control (T₁₆). The treatment with NAA 50 mg/L seed hardening + spraying at 30 DAS (T₁₅) recorded significantly highest root dry matter accumulation per plant (1.582, 1.885 and 1.734 g) at 45 DAS, (2.369, 2.772 and 2.570 g) at 60 DAS and (2.673, 3.073 and 2.873 g) at harvest during 2016, 2017 and in pooled analysis, respectively. While untreated absolute control (1.102, 1.409 and 1.256 g) at 45 DAS and at 60 DAS (1.739, 2.139 and 1.939 g, respectively) recorded significantly the lowest root dry weight per plant (Table 3). The increase in root dry weight may be due to enhanced lipid utilization through glyoxalate cycle, a primitive pathway leading to faster growth and development of root by increasing cell division and cell enlargement and thereby enabling them to produce relatively more quantity of dry matter. Increase in root dry weight by seed hardening was also recorded by Prakash et al., (2013) in rice and Gurudev Singh et al., (1991) in wheat and Kinjal (2017) in Urd bean. There was a less rate of increase in the root dry matter after 60 DAS from pod development stage till the harvest leading to an increase in pod weight.
 
Reproductive dry matter accumulation at various growth stages
 
In green gram crop, the flowering i.e. reproductive phase starts after 35 DAS. Therefore, the reproductive dry matter production per plant was increased progressively with the advancement of growth stages from 45 days after sowing till harvest of the crop and all the treatments differed significantly. The significant differences were occurred for the reproductive dry matter accumulation per plant at 45 days after sowing. The treatment with CaCl₂ 2% seed hardening + 1% spraying at 30 DAS (T₁₁) recorded significantly the highest (1.903, 2.317 and 2.110 g) at 45 DAS, at 60 DAS (10.100, 12.013 and 11.057 g) and (14.713, 17.213 and 15.963 g) at harvest during 2016, 2017 and in pooled analysis, respectively as compared to untreated absolute control (Table 4).
 
Total dry matter partitioning at various growth stages
 
The total dry matter production per plant is aggregate of stem, leaf, root and reproductive dry matter accumulation at respective growth stages. It was increased progressively and differed significantly at all the growth stages from sowing to harvest in green gram. The results pertaining to the total dry matter (TDM) production and partitioning indicated significant differences and revealed that pre-sowing seed hardening with 2% CaCl₂ (T₁₁ and T₁) recorded highest total dry weight per plant (4.393, 5.043, 4.718 and 4.383, 5.000, 4.692 g) while lowest total dry matter production was recorded (3.450, 4.033 and 3.742 g) in untreated absolute control (T₁₆) at 30 DAS during summer 2016, 2017 seasons and in pooled analysis, respectively. The treatment with CaCl₂ 2% seed hardening + 1% spraying at 30 DAS (T₁₁) recorded highest total dry matter accumulation per plant (13.013, 14.537 and 13.775 g) at 45 DAS, at 60 DAS (27.057, 30.417 and 28.737 g) and at harvest (29.720, 33.420 and 31.570 g) during summer 2016, 2017 seasons and in pooled analysis, respectively (Table 5). The increase in TDM production and partitioning towards maturity may be due to indeterminate growth pattern, higher rate of CO₂ fixation and RuBP carboxylase activity during crop growth. The plant growth regulators were more effective in increasing the TDM production as compared to seed hardening treatments. The association of TDM partitioning with grain yield was more significant at all the stages of crop growth. Thus, TDM production and partitioning is an important parameter in boosting the source-sink relationship and ultimately yield potential. Similar observations were also made by Nam et al., (1998), Katti et al., (1999), Manjunatha (2007) and Sujatha (2014) in chickpea. Thus, the total dry matter is composed of more with the leaf and stem dry weight and little with root dry weight during vegetative growth phase. But during reproductive growth phase, reproductive parts like pods, flowers contributes more as compared to leaf, stem and root dry weight. Murthy et al., (2002) corroborated the increase in dry matter in any genotype might reflect in increase in yield contributing characters and ultimately final economic yield and positive correlation of dry pod yield with dry matter accumulation.
 
Seed yield (kg ha^-1) and harvestindex (%) at harvest
 
The data on seed yield per hectare and harvest index at harvest is reported in Table 6. The significantly highest seed yield per hectare (949, 1006 and 978 kg ha^-1) was recorded by the treatment CaCl₂ 2% seed hardening + 1% spraying at 30 DAS (T₁₁) and remained at par with the treatments of CCC 1000 mg/L seed hardening + spraying at 30 DAS (T₁₃) (922, 964 and 943 kg ha^-1) and  NAA 50 mg/L seed hardening + spraying at 30 DAS (T₁₅) (893, 917 and 905 kg ha^-1) while significantly the lowest seed yield per hectare was observed in the untreated absolute control (639, 679 and 659 kg ha^-1) during 2016, 2017 and in pooled analysis, respectively. The harvest index was non significant during both the years but differed significantly among the treatments in pooled analysis. The treatment T₁₁ recorded significantly highest harvest index (30.15%) in pooled analysis and remained at par with the treatments T₁₃ (30.01%), T₁₅ (29.84%), T₁₂ (29.49%), T₁₄ (29.27%), T₆ (28.80%), T₈ (28.66), T₁₀ (28.35%) and T₇ (28.34%). While, the treatment of absolute control (T₁₆) recorded significantly the lowest (26.84%) harvest index.


 
Improvement in yield according to Humphries (1979) could happen in two ways i.e., by adopting the existing varieties to grow better in their environment or by altering the relative proportion of different plant parts so as to increase the yield of economically important parts. The influence of plant growth regulators and seed hardening chemicals significantly increased the seed yield. The increased seed yield could be attributed to higher dry matter production and its accumulation in reproductive parts. This could probably be due to enhancement of photosynthesis and nitrogen metabolism which are the major physiological process influencing plant growth and development, better carbon assimilation, better accumulation of carbohydrates and reduced respiration in plants. These results are in agreement with the findings of Mahabir Singh and Rajodia (1989) in soybean, Singh and Dohare (1964), Das and Prusty (1982) and Pothalkar (2007) in pigeon pea. The harvest index (HI) is the best measure of source-sink relationship indicating the efficiency of genotype to convert biological yield into economic yield.

The leaf dry matter decreased in all the treatments after 60 DAS at the time of pod development upto harvest. There was a very less rate of increase in the stem and root dry matter from pod development stage after 60 DAS till the harvest leading to an increase in pod weight. This suggests the possible translocation of stored carbohydrates from leaves, stem and root to reproductive parts for pod development. Particularly the translocation process must have been more prominent in the treatment CaCl₂ 2% seed hardening + 1% spraying at 30 DAS (T₁₁) than rest of the treatments and was very less in untreated control treatment indicating that photosynthesis and translocation efficiency increases by seed hardening and foliar spraying treatments with agrochemical and growth regulators for increasing the pod yield. Therefore, the treatment T₁₁ was more efficient in dry matter production and its partitioning in leaves, stem, root and pods by allocating maximum dry matter to the pods. The influence of plant growth regulators and seed hardening chemicals significantly increased the seed yield and harvest index.