Source of seeds
Genetically pure delinted and fuzzy seeds of American varieties (H 1300 and H 1098-i) of cotton were procured from Cotton Section, Department of Genetics and Plant Breeding, CCS HAU Hisar. All aspects of the experimental research and field studies on plants, including the collecting of plant material, were carried out in a manner that was in accordance with the applicable institutional, national, and international norms and legislation.
Source of nanoparticles
Zinc oxide nanoparticles (ZnONPs) and titanium dioxide nanoparticles (TiO2NPs) were synthesized in laboratory of Department of Seed Science and Technology and Centre of Nanotechnology, CCS HAU Hisar by chemical method of synthesis using Zn(NO3)2.4H2O and titanium tetra isopropoxide, respectively.
Synthesis procedure of ZnONPs and TiO2NPs and characterization of nanoparticles were described in my research paper i.e., Singh et al., [10].
Seed treatment
Seeds were nanoprimed with nanoparticles for 10 hours and seeds which were only hydroprimed for 10 hours were considered as control in experiment. The treated seed were stored under ambient condition in plastic bags.
Observations recorded
Seed quality parameters i.e., moisture content (%), germination percentage, seedling length (cm), seedling dry weight (mg). Seed vigour index-I, seed vigour index-II, dehydrogenase activity and electrical conductivity were recorded at different time interval (0, 3, 6, 9 and 12 months) during storage. During storage, observed temperature and relative humidity is mentioned in Table 3.1.
Table 3.1
Meteorological data on temperature and relative humidity during the course of investigation
Month | Average temperature (oC) | Relative humidity |
---|
Maximum | Minimum | Morning | Evening |
---|
March 2019 | 26.5 | 10.5 | 87 | 42 |
April 2019 | 36.7 | 18.4 | 69 | 27 |
May 2019 | 39.0 | 21.6 | 59 | 26 |
June 2019 | 40.5 | 25.8 | 68 | 33 |
July 2019 | 35.4 | 25.7 | 82 | 63 |
August 2019 | 34.7 | 26.1 | 86 | 63 |
September 2019 | 35.3 | 26.0 | 86 | 55 |
October 2019 | 32.6 | 17.9 | 85 | 38 |
November 2019 | 26.9 | 12.9 | 89 | 46 |
December 2019 | 17.1 | 5.7 | 94 | 68 |
January 2020 | 17.1 | 5.2 | 97 | 65 |
February 2020 | 22.7 | 6.8 | 93 | 51 |
March 2020 | 25.9 | 12.4 | 92 | 56 |
Seed moisture content (%):
Moisture content of seeds was calculated according to ISTA (International Seed Testing Association) by hot air oven method. Ten grams of seeds from each lot were kept in moisture box in the oven at 103 ± 1°C for 24 hours. The samples were taken out and cooled before weighing. Moisture content was calculated as given below:
$$\text{M}\text{o}\text{i}\text{s}\text{t}\text{u}\text{r}\text{e} \text{c}\text{o}\text{n}\text{t}\text{e}\text{n}\text{t}\left(\text{%}\right)=\frac{\text{M}2-\text{M}3}{\text{M}2-\text{M}1}\times 100$$
Where,
M1 = Weight of moisture box alone
M2 = Weight of moisture box and sample before drying
M3 = Weight of moisture box and sample after drying
Standard germination (%)
Germination test was conducted using three replicates by adopting “Between paper method” as described by ISTA. At the final count of germination test, the number of normal seedlings in each replication was counted and the germination was calculated and expressed in percentage.
Seedling length (cm)
Ten normal seedlings in three replications of each variety were selected randomly during germination test and measured in centimeter and average seedling length was calculated.
Seedling dry weight (mg)
Ten normal seedlings used for measuring root and shoot length were kept in butter paper and dried in a hot-air oven maintained at 75 ± 10C temperature for 24 hours. Then the seedlings were removed and allowed to cool in a desiccator for 30 minutes before weighing on an electronic balance. The average weight was calculated and expressed as dry weight of seedlings in milligrams.
Seedling vigour indices
The seedling vigour indices was computed using the formula as suggested by Abdul-Baki and Anderson [11] and expressed as whole number.
Seedling vigour index-I = Standard germination (%) × Average seedling length (cm)
Seedling vigour index-II = Standard germination (%) × Average seedling dry weight (mg)
Electrical conductivity (µS/cm/seeds):
Electrical conductivity of the seed leachates was measured to know the status of membrane permeability as per ISTA [12]. For this, 50 seeds selected randomly replicated thrice from each seed lot were soaked in separate beakers each containing 75 ml of distilled water. The seeds were immersed completely in water and beakers were covered with the foil. Thereafter, these samples were kept in the germinator at 25 + 1°C for 24 h. The electrical conductivity of seed leachates was measured by 60 direct reading conductivity meter. The conductivity was expressed in µS/cm/seeds.
Dehydrogenase activity test:
Dehydrogenase activity test [13], reduction of 2,3,5-triphenyl tetrazolium chloride to red formazan by dehydrogenase enzyme in seed embryo is the basic principle for topographical tetrazolium test for seed viability. It is a quantitative method, which may be used to determine varying dehydrogenase activity between seeds of similar viability and therefore, it is a measure of seed vigour. Sample of 1 g of each treatment was grounded to pass through a 20-mesh screen to obtain 200 mg flour. The flour was soaked in 5 ml of 0.5% tetrazolium solution at 38 oC and was centrifuged after 3–4 h at 10,000 rpm for 3 min and supernatant was poured off. Formazan was extracted with 10 ml acetone for 16 h followed by centrifugation and then absorbance of the solution was determined by Systronic spectrophotometer 169 at 520 nm wavelength.