Ecient Use of Nitrogen, Gibberellic Acid and Potassium on Canola Production under Sub-tropical Regions

The global demand for crop production is rapidly growing due to the continued rise in world population. Crop productivity varies generally with soil nutrient prole and climate. The optimal use of fertilizers might help to attain higher crop yield in canola. To circumvent nutrient imbalance issues in soil, two separate eld trials were conducted to determine the best source of nitrogen (ammonium sulfate/ammonium nitrate), foliar application of gibberellic acid (GA 3 ) and potassium (K) for the canola yield and yield attributes for four consecutive years (2014 to 2018). Both experiments were carried out in randomized complete block design (RCBD) with three replicates. The nitrogen source in the form of ammonium sulfate (0, 10, 20 and 30 kg/ha) and ammonium nitrate (0, 50, 75 and 100 kg/ha) was applied in the rhizosphere. In another experiment, the canola crop was sprayed with four level of gibberellic acid (GA 3 ; 0, 10, 15, 30g/ha) and potassium (K; 0, 2.5, 3.5, 6g/ha) separately or in combination by using hydraulic spryer. The analysis showed that fertilization with nitrogen in the form of ammonium nitrate (100 kg/ha) and ammonium sulfate (30 kg/ha) had a positive effect on the plant height, number of branches, fruiting zone, seed yield per plant, seed yield per hectare of canola except oil percentage. Moreover, canola plants displayed a signicant improvement on all studied features with high dosses of GA 3 (30 g/ha) and K (6 g/ha) individualy and in combined form. In a nutshell, compared to both source of nitrogen, ammonium nitrate was more ecient and readily available source of nitrogen. Gibbrellic acid being a natural growth elicitor and potassium as a micronutrient serve as potential source to improve yield and to mange nutrient prole of canola.


Introduction
For several years now, Arid and semi-arid areas located in certain third world countries have been facing massive shortage of edible oils which was met through imports in large quantities from other countries 1 .
As a result, efforts aiming at reducing the imbalance between the production and consumption for edible oils have been made by under-developed countries. In this context, oil seed crops seemed to be an accurate option for these countries. Among these crops, canola appeared as a potential candidate for the domestic edible oil production 2 . This could be explained by to the low content of erucic acid and glucosinolates in oil and seed cake, respectively 3 . Moreover, canola crop can survive under diverse environmental conditions due to a wide range of adaptability 4 . However, mismanagement and highly imbalanced application of micro and macronutrients found to be reducing the yield of canola crop, therefore, nutrients management strategies for optimizing the canola production are highly required 5 Balanced fertilizer application in uence the crop yield, quality and the soil productivity 6 . The adequate nitrogen supply is important in order to boost up the canola productivity and it holds a key role in plant tissue growth and development. Plus, it represents a part of chlorophyll, nucleotides, protein, and amino acids formation which directly affect the quality and quantitative traits of the crop. Other factors such as Soil pro le, texture, and moisture content uctuation at various critical stages of growth and development of canola may in uence the nitrogen use e ciency on canola crop. Actually, this kind of crop can very responsive to fertilizer application, especially nitrogen which signi cantly impacts the plant height, number of branches/plant, number of ower/plant, number of pods/ plant and their weights, and seed yield/ ha. It also effects of the leaf area (LA) development and LA duration after owering in canola crop 7,8 Many natural and arti cial plant regulators may be used with the aim of controlling the developmental process from germination to post-harvest preservation of crop plants and subsequently, optimizing their production 9 . Among these fertilizers, gibberellic acid (GA 3 ) is obviously a key regulator product for plantgrowth and other physiological mechanisms. It can stimulates the root and stem elongation, seed germination ,break dormancy, leaf expansion, fruit senescence, and owering 10 . Moreover, GAs may in uence the metabolic pathways including nitrogen metabolism, chlorophyll production and degradation, nitrogen redistribution, and translocation of assimilates 11 . It can also induces the expression of several hydrolytic enzymes involved in the conversion of starch to sugar which ultimately in uence the plant growth at vegetative and reproductive stages 12 , plant signaling mechanisms, gene expression, and plant morphology and physiology 13,14 .
Besides nitrogen and phosphorus fertilizers, potassium (K) found to be in uencing the seed oil content percentage, yield and yield-contributing traits of the canola crop 15,16 . K is very important fertilizer which is involved in photosynthesis, regulation of stomata, control of the ionic balance, translocation of photosynthates, protein synthesis, enzymatic activities, and many other physiological and biochemical processes 17,18 . Therefore, K is considered as primary osmoticum that plays an important role at maintaining the low water potential in plant tissue and also impacts the plant growth and development.
For plant breeders, the e cient use of nutrients from the soil by the crop plants is a promising characteristic. Some plants may produce high yields with minimal inputs 14,18 . Many studies showed that signi cant variation exists among various genotypes of canola regarding e cient use of potassium 19 .
Keeping in view the possible outcomes of e cient use of K and GA, the current study evaluates canola genotype (Surhan-2012) for four consecutive years for these traits. Hence, current manuscript demonstrated the in uence of foliar application of GA 3 and K separately, or in combination in canola.
This study carries immense importance as a reference for the impact of these two important nutrients on canola production and the multi-year screening of Surhan-2012 in this context.

Experimental Design
The eld experiment was conducted at the farm of Nuclear Institute of Agriculture (NIA), Tando Jam, Sind, Pakistan (31° 25' 0" North, 73° 5' 0" East) during the four seasons of 2014-2018. Data were collected under a randomized Complete Block Design (RCBD) with three replications per block.
The canola seeds were collected from nuclear institute of Agriculture (NIA) TandoJam and sown. The plants thinned after 15-20 days of germination for the purposes of maintaining long distance dispersal of plants. All the recommended agronomic and cultural practices that govern the production of the crop were applied e ciently during the plant growth cycle 20 .

Application of ammonium sulfate [(NH 4 ) 2 SO 4 ] and ammonium nitrate [(NH 4 ) NO 3 ] as Nitrogen supplements
The four levels of ammonium sulfate (0, 10, 20 and 30 kg/ha) and ammonium nitrate (0, 50, 75 and 100 kg/ha) were used as nitrogen source. Both nitrogen fertilizers were applied in two split doses; the rst dose was applied after three weeks of crop sowing whereas the second was undertaken after seven weeks of sowing. One square meter (m 2 ) area of plants was chosen randomly from each plot for harvesting during four seasons of 2014-18. The agronomic parameters of crops were computed from the plant height (cm), number of branches/plant, fruiting zone length (cm), seed yield/plant (g), seed yield/ha (kg). the differences of oil content (%) of canola seeds were recorded, pooled and statistically analyzed in order to evaluate the effect of different sources/doses of nitrogen on the agronomic characters and traits of canola 16 .
The effect of potassium nitrate and Gibberellic Acid on canola seed germination Ten different combinations (Table 1) of K and GA 3 were applied as foliar spray. The experiment was carried out using a randomized complete block design (RCBD) with three replications.
Before the foliar application, GA 3 was dissolved in ethanol and mixed with water. Various dilutions were then made in order to obtain solutions with several concentrations. The different combinations of GA 3 and potassium were sprayed after one month of sowing. The treatments were applied three times at one week intervals and the control plants were sprayed with distilled water only. One m 2 area of plants was chosen randomly from each plot at harvesting time during four seasons (2014-18). The data of agronomic parameters including Plant height (cm), number of branches/plant, fruiting zone length (cm), seed yield/plant (g), seed yield/ha (kg) and oil content percentage have been recorded according to the protocol reported by A.O.A.C in 1980. Subsequently, the recorded data were analyzed using analysis of variance (ANOVA) combined with HSD (Steel et al., 1997). Tukey's test was also used to determine the signi cant difference between the treatments with the help of statistical software SAS (version 9.4) and nally calculation of the Cost-bene t ratio.

Results And Discussion
Effect of ammonium sulfate (NH 4 ) 2 SO 4 as nitrogen source on crop production The statistical analyses performed during all four seasons showed signi cant differences on all studied features (  Table 3). Another important factor that must be taken into account is the Sulfur element since it plays an important role in the synthesis of proteins which in turn affects the oil contents in canola seeds 21 . Hence, the balanced application of S and N is vital with the objective of further improving the canola seeds quality and production 21 . Karamanoset al. 22 suggested that the optimal ratio of N:S ranging from 7:1 to 5:1 can maximize canola production. In fact, a study conducted by Brennan and M.D.A, 2008 and proven that the canola production can be extremely limited in case of sulfur de ciency in soil 23 . The supply of arti cial sulfur promotes the nitrogen uptake e ciency of canola production and consequently elevates the level of protein in leaves: this will de nitely enhance the crop productivity and yield 24 .
Our results are in agreement with those reported by Chienet al. 25 (Table 3). From these ndings it can be concluded that a nutrient de ciency (nitrogen) can severely hampers canola productivity 28, 29 . Furthermore, the canola yields can be enhanced by a better management of nitrogen at the optimum growth stages of canola 2,16 . Nitrogen is an essential plant nutrient that simulates its meristematic activity, cell elongation, and elevates the photosynthesis of canola. These factors will ultimately boost growth and yield of the canola plant 30 31 . Similar ndings have been made in other studies highlighting the importance of nitrogen supplementation in the re nement of the rapeseed yields in diverse agro-climatic conditions 32 .
As far as we know, Nitrogen has strongly and signi cantly correlated with the seed yield per hectare, plant height, number of branches per plant, fruiting zone length, and number of seed per plant, in addition to the enhancement of the number of pods per seed, 1000 seed weight, biological yield, seed yield , and oil yield 27,32 . On this basis, it can be concluded that the canola production depends on the selection of the correct dose, source, and timing of nitrogen fertilizer application. Unbalanced application of nitrogen fertilizer may adversely affect the canola production 6 . The source of nitrogen fertilizer may also change the plant N uptake and soil N availability and hence impacting the ultimate canola productivity 33 . In our experiments, two sources of nitrogen were tested and compared one with another. The subsequent results showed that ammonium nitrate had signi cantly contributed to the enhancement of canola production comparing to the ammonium sulfate 34 . It has been reported elsewhere that the application of ammonium sulfate reduces the pH of the soil as well as dissolution of many other nutrients resulting in negative impacts on plant growth and development compared to ammonium nitrate 35 .
Effect of foliar application of Gibberellic acid and potassium fertilizer on canola yield and yield components The in uence of various treatments related to the application of gibberellic acid and potassium fertilizer were also studied in accordance with the yield parameters of canola. The results of the present study provide evidence that all the agronomic traits and oil percentage tend to increase with increasing levels of foliar application of K and GA 3 solely or combined in comparaison with the unsprayed plants. A signi cant increase was recorded using different treatements of GA3 and K in plant height, number of branches per plant, fruiting zone length, seed yield per plant, seed yield/ha and seed oil percentage compared to control (T 1 -T 10 ).
The measurement values of plant-height in all of the treatments were higher than the control plant during a four-year period (2014 to 2018). Signi cant differences were also observed among the treatments (F=81.913; p≤ 0.0000, F=99.79; p≤ 0.0000, F=86.782; p≤ 0.0000, and F=101.34; p≤ 0.0000) during all seasons ( Table 4). The maximum plant-height was reported when combined GA 3 (30kg/ha) and K (6 g/m 2 ) ( T 10 followed by T 9 , T 8 and so on) ( Table 4) were applied. However, both T 4 (GA 3 0 and K 6.0) and T 8 (GA 3 10kg/ha and K 2.0g/m 2 ) showed an almost insigni cant variation in the plant-height meseaurements compared to other treatments.
The foliar application of K and GA 3 signi cantly affected the number of branches per canola plant comparing to the control one (T 1 ). The highest number of branches per plant were recorded in T 10 (30GA 3 g/ha+6.0 g/m 2 K) which appeared to have the same trend as that reported for canola plant-height measurements (Table 5). A considerable rise in the fruiting zone length (cm) was also observed when combined foliar applications were applied (T 10 ). The signi cant differences among the treatments (F=101.814; p≤ 0.0000, F=123.32; p≤ 0.0000, F=126.62; p≤ 0.0000 and F=122.4; p≤ 0.0000) were also noted for over four years of the study (Table 6). As Another agronomic trait was affected when foliar applications of K and GA 3 were applied ( individually or combined) is the number of seeds per plant: it was found that canola plants produce more number of seeds per plant when combined GA 3 and K were applied (T 10 ) during the four seasons of 2014-2018 (Table 7). This parameter seemed to be improved immeasurably in all treatments (T 2 -T 10 ) compared to the control plant (T 1 ). Therefore, it can be concluded that improvement of this agronomic parameter can be successfully attained when increased rates of the foliar (K and GA3) were applied.
Since seed yield ha -1 is considered as the main interest for canola breeders, higher rates of the foliar (K and GA 3 ) were applied (individually or combined) with respect to non-sprayed plants (T 1 ); the obtained results of this investigation indicate that high seed yield occurs in all treatments (T 2 -T 10 ), particularly, when increased rates of the foliar were applicable.
The changes in oil percentages, in response to K and GA 3 application were also investigated. The highest oil percentage was observed at the T 10 treatment followed by T 3 and T 5 during the four cropping seasons of 2014-18 (Table 9).
In view of the aforementioned ndings, it can be concluded that combined form of GA 3 and K (T 10 ) presents a potential strategy to enhance growth performance of canola. The promoting effect of gibberellic acid and potassium treatments contribute to the metabolic and other physiological processes leading to better crop yields. Interestingly, for the majority of the studied traits, the K application (T 4 ) acts similarly and almost insigni cantly to the combined application (T 8 ) of K (3.5g/m 2 ) and GA 3 (15g/ha), this could be attributed to the key role of K in improving canola yields (Table 4-9).
GA 3 and K fertilizer application is necessary to increase the vegetative and reproductive growth of canola plant 36 . These fertilizers could be involved in improving defence mechanisms of canola plant which may consequenlty affect the seed yield. Similar results have been reported using these same treatments on sesame plant 28 . Likewise, foliar application of potassium and gibberelic acid alone or in combination incerases the plant vegetative and reproductive growth of the plant resulting in the enhancement of the yield per unit 37 . In fact, in a study reported by Imran and A.A. Khan, 2017, the application of K fertilizer not only enhances the yield per unit, fresh nut and karnel dry mass (spliting percentage), it also reduces the blank percentage 38 . It was also observed that in absence of gibberellic acid applications, the blank percentage and spliting percentage could be ameliorated 39 .
Jan et al (2019) reported that high concentrations of potassium K and Zing Zn after the simultaneous foliar applications of GA 3 and K separately or in combination could be found in canola plant leaves 40 .
The evidences of this study suggest that the interactive effects of GA 3 and K can be employed in the aim of improving morphological aspects and yield attributes of canola. It can also be expected that these interactive effects may elevate the plant resistance against various biotic and abiotic stresses, carbohydrate translocation,and the photosynthesis process 39 . Khan et al (2019) also mentioned that these fertlizers (GA 3 and K) might strengthen the defence mechanism of the plant which ultimately impacts the plant growth and yield 41,42 . In short, with appropriate application of nitrogen fertilizers, GA 3 , and K, canola yeilds can be substantially improved.

Ethic statements
This study does not involve any wild or endangered species of plants. Moreover, it does not encompass the collection of any new plant material. The study is only related to the eld performance of a canola genotype under different growth treatments. The seeds of canola were collected from nuclear institute of Agriculture (NIA) tandojam. The evaluated genotype is a cultivated species and is a released variety in Pakistan. The experiment was conducted in line with institutional and national policies.

Conclusion
Nitrogen is a an essentiel nutrient for the metabolic function and production process of the canola plant or any other plant. Therefore, the canola yields can be monitored with the application of nitrogen fertilizers pertaining to different sources and proportions. The optimum levels of nitrogen fertilizer were found to be 30 kg/ha ammonium sulfate and 100 kg/ha ammonium nitrate. These data have been obtained according to the of agronomic yields of a four-year study (2014-18). Another fact to consider is that Ammonium nitrate (NH 4 NO 3 ) is more e cient and readily available source of nitrogen compared to ammonium sulfate [(NH4) 2 NO 3 ]. This study has recommended the optimum value and source in subtropical region of the world. On the contrary, gibberellic acid and potassium in uence the plant growth and its development, enable the plant to survive in nutrient de cient soil and increase the yield in the four growing seasons (2014-18). It is suggested that canola plant illustrated maximum potential of yield at high dose of GA 3 (30g/ha) andK (6.0 g/m 2 ) alone or in combination.