DESC:
SUMMARY OF THE INVENTION
One of the aspects of the present invention provides a rapid detection kit (100) for the viability of seeds, the kit (100) comprising:
i. a base plate (101) with seed wells;
ii. a seed puncher/crusher (102);
iii. a reagent A (103); and
iv. a reagent B (104);
wherein the base plate provides seed wells to place the individual seed;
wherein seed puncher presses the seed to squeeze the seed content out into the well;
wherein reagent A (103) acts as an extractant;
wherein reagent B (104) reacts with the seed extract to distinguish between viable and non-viable seeds; and
wherein the said kit provides seed viability by indicating colour change.
Another aspect of the present invention provides a method for determining seed viability, the method comprising:
i. crushing the seeds to obtain a seed powder;
ii. contacting the seed powder with Reagent A for a time ranging from 10 minutes to 45 minutes at an ambient temperature to obtain a seed extract;
iii. filtering the seed extract to obtain a filtrate and a residue comprising suspended particles of the seeds;
iv. combining the filtrate with Reagent B to obtain a reaction mixture;
v. incubating the reaction mixture in a light chamber at light intensity in the range from 20000 lux to 30000 lux.for a time ranging from 2 minutes to 10 minutes; and
vi. observing the colour change of the reaction mixture from colourless to pale yellow or pink-reddish or pink or red to determine the seed viability;
wherein colourless or pale yellow colour indicates non-viable seeds, pink reddish, pink colour to red colour indicates at least 80% viable seeds.

BRIEF DESCRIPTION OF THE FIGURES
Figure 1 shows different components of the kit (100)
Figure 2 depicts Lot 3 testing
Figure 3 shows results of the Seed Testing with rapid detection method
Figure 4 depicts the outcome of Seed Testing with conventional germination method:
Figure 5 shows Spectrophotometric analysis of the sample depicting the clear pattern between viable and non-viable seeds.
Figure 6 show Light incubation chamber

DETAILED DESCRIPTION OF THE INVENTION
In the specifications, different types of terms are used for understanding purpose. It does not limit the scope of the present invention. The definitions of the terms are provided below:

Definitions:
The term ‘seed(s)’ or ‘onion seed’ used herein refers to seed is the small, hard part of a plant from which a new plant grows. The seed(s) may include but not limited to vegetable seeds, fruits seeds, spices seeds, grains or grains seeds, onion, rice, wheat, maize, sorghum, barley, peanut, soybean, lentil, common pea, common bean, walnut, pecan, and sunflower. The term ‘seed(s)’ can be used as singular or plural form.
Terms ‘seed’, ‘seed(s)’ ‘seed(s)’ or ‘onion seed’ can be used interchangeably in the specification.
The term ‘seed testing’ used herein refers to a test that determine the standards of a seed lot viz., physical purity, moisture, germination and ODV and thereby enabling the farming community to get quality seeds.
The term ‘viability counting’ or ‘viability testing’ used herein refers to a method used in to determine the seeds that develop into seedlings.
The term ‘material’ used herein refers to a substance that can be used for making of kit or an apparatus. The material may include but not limited to clay, cardboard, hydrogels, polymers (natural/ synthetic) with additives like fertilizers, plant growth promoters, fungicides, pesticides, (organic, natural, synthetic) and/or combinations thereof.
The present invention relates to a method of improving germination by subjecting seeds to pre-sowing detection method. More particularly, it relates to a rapid detection kit for onion viability technology to be applied to seeds prior to sowing for the purpose of achieving rapid and uniform germination of seeds after sowing and improving the germinating rate even in an adverse environment.
Once sowed, seeds germinate after a certain period of time if such environmental conditions as moisture, temperature, oxygen and light are all appropriate. During this pre-germination period from sowing to germination, various metabolic processes proceed within seeds. The length of said period varies depending on the degree of maturity of each individual seed, the kind of seed, and such environmental factors as temperature and moisture content. Shortening of said pre-germination and attaining rapid and uniform germination will facilitate cultivation management and, in commercial production, offer such advantages as reduction in production costs and reduction of labour.
The present invention provided a new rapid germination kit for viability of onion seeds for germination, wherein the kit uses novel reagents that increases the accuracy, simplify the testing procedure and reduces the testing time of seed viability testing.
A rapid detection kit (100) for the viability of seeds, the kit (100) comprising:
i. a base plate (101) with seed wells;
ii. a seed puncher/crusher (102);
iii. a reagent A (103); and
iv. a reagent B (104);
wherein the base plate provides seed wells to place the individual seed;
wherein seed puncher presses the seed to squeeze the seed content out into the well;
wherein reagent A (103) acts as an extractant;
wherein reagent B (104) reacts with the seed extract to distinguish between viable and non-viable seeds; and
wherein the said kit provides seed viability by indicating colour change.
One of the embodiments of the present invention provides a rapid detection kit for the viability of seeds, the kit (100) comprising:
i. a base plate(101) with seed wells;
ii. a seed puncher/crusher (102);
iii. a reagent A (103); and
iv. a reagent B (104)
Another embodiments of the present invention provides a rapid detection kit for the viability of onion seeds, the kit (100) comprising:
i. a base plate(101) with seed wells;
ii. a seed puncher/crusher (102);
iii. a reagent A (103); and
iv. a reagent B (104)
Another aspect of the present invention provides a method for determining seed viability, the method comprising:
i. crushing the seeds to obtain a seed powder;
ii. contacting the seed powder with Reagent A for a time ranging from 10 minutes to 45 minutes at an ambient temperature to obtain a seed extract;
iii. filtering the seed extract to obtain a filtrate and a residue comprising suspended particles of the seeds;
iv. combining the filtrate with Reagent B to obtain a reaction mixture;
v. incubating the reaction mixture in a light chamber at light intensity in the range from 20000 lux to 30000 lux.for a time ranging from 2 minutes to 10 minutes; and
vi. observing the colour change of the reaction mixture from colourless to pale yellow or pink reddish or pink or red to determine the seed viability;
wherein colourless or pale yellow colour indicates non-viable seeds, pink reddish or pink colour to red colour indicates at least 80% viable seeds.
Another embodiment of the present invention provides a method for determining seed viability by using the rapid detection kit, the method comprising:
i. putting seed(s) in the wells (one seed per well) of a baseplate (101) of the kit (100);
ii. pressing a seed crusher (102) on the seeds in the well to squeeze out the seed content;
iii. adding 100ul a reagent A (103) to each well and wait for time period in the range of 60 seconds to 600 seconds;
iv. adding 50ul a reagent B (104) to each well and incubating the plate in light chamber for 3 min and observing colour change from colourless to pale yellow to pink reddish;
v. identifying and counting the number of pink reddish wells corresponding to viable seeds; and
vi. Calculating the seed viability.
Yet another embodiment of the present invention provides the rapid detection kit for the viability of seeds, wherein the kit (100) is placed in a light incubation chamber (105) for suitable time to incubate the reaction mixture.
Another embodiment of the present invention provides a rapid detection kit for the viability of onion seeds, wherein the base plate (101) can be made up of non-reacting materials including such as but is not limited to glass, acrylic, teflon, etc.
Another embodiment of the present invention provides a rapid detection kit for the viability of onion seeds, wherein the base plate (101) can be of any dimensions based on the quantity of seeds to be measure. The base plate has wells with depth in the range of 0.5cm to 2cm and diameter in the range of 0.5cm to 2cm and these dimensions can vary depending on the seed type. The number of well can be 50, 100, 200 or in the multiple of 50. The wells are used to place the individual seed.
Yet another embodiment of the present invention provides a rapid detection kit for the viability of onion seeds, wherein seed puncher/crusher (102) can be made up of materials including such as but is not limited to SS316, SS304, and Teflon etc.
Further embodiment of the present invention provides a rapid detection kit for the viability of onion seeds, wherein the seed puncher consists of teeth like protruding makeup in same number as base plate well. The function of puncher is to press the seed to squeeze the content out into the well.
Other embodiment of the present invention provides the method for determining seed viability, wherein the seeds are washed and dried before crushing.
Further embodiment of the present invention provides the method for determining seed viability, wherein the seed powder is having particle size less than 400 micron.
Another embodiment of the present invention provides the method for determining seed viability, wherein the Reagent A is a solvents selected from water, methanol, ethanol, propanol, ethyl acetate, hexane, and combination thereof. Furthermore, the reagent A (103) may include a polymer and can be organic or inorganic form the below group including such as but is not limited to acrylamide, cellulose, ulvan, etc.
In an embodiment, reagent A (103) acts as an extractant.
Yet another embodiment of the present invention provides the method for determining seed viability, wherein the ratio of seed powder to the Reagent A is 1:10.
Another embodiment of the present invention provides a rapid detection kit for the viability of onion seeds, wherein reagent B (104) may be specially designed compounds belonging to the following group of chemicals including such as but is not limited to azo compound or halogen compound or metal nitrates.
In an embodiment, reagent B (103) is responsible for a reaction that distinguishes between viable and non-viable seeds.
Another embodiment of the present invention provides the method for determining seed viability, wherein the reagent B is azo compound, selected from Azobenzene, P-Azobenzenearsonate, 1,1'-Azobis-1,2,3-triazole, 2,2'-Azobis(2-amidinopropane) dihydrochloride, 4,4'-Azobis(4-cyanopentanoic acid), Azobisisobutyronitrile, Azodicarbonamide, Azomethane, Azoxy compounds, Para-Azoxyanisole, Azoxybenzene, Balsalazide, Diethyl azodicarboxylate, Diimide, Diisopropyl azodicarboxylate, 4,4'-Dinitro-3,3'-diazenofuroxan, 1,3-Diphenyltriazene, Calmagite, Fazadinium bromide, Glycoazodyes, Methylazoxymethanol, Methylazoxymethanol acetate, Phenazopyridine, 3-Phenylazoacetylacetone, Pigment Red 178, Potassium azodicarboxylate, Tetramethylazodicarboxamide, Tetrazene explosive, Thorin (chemistry), Toluidine red.
Another embodiment of the present invention provides the method for determining seed viability, wherein the reagent B is halogen compound selected from hydrofluoric acid (HF),sodium fluoride (NaF), hydrochloric acid (HCl) and sodium chloride (NaCl), chloroform (CHCl3), carbon tetrachloride (CCl4), bromine water hydrobromic acid, iodine (I2), potassium iodide (KI), iodine pentafluoride, astatine monochloride (AtCl), chlorine dioxide (ClO2), bromine chloride (BrCl), and iodine trichloride (ICl3), fluoromethane (CH3F) and chlorofluorocarbons (CFCs), perfluorooctanoic acid (PFOA)
Other embodiment of the present invention provides the method for determining seed viability, wherein the reagent B is metal nitrates selected from Barium nitrate, Cesium nitrate, Lithium nitrate, Potassium nitrate, Rubidium nitrate, Silver nitrate, Sodium nitrate.
Further embodiment of the present invention provides the method for determining seed viability, wherein the reagent B is preferably silver nitrate.
Another embodiment of the present invention provides the method for determining seed viability, wherein the concentration of reagent B ranges from 10mM to 200mM.
Yet another embodiment of the present invention provides the method for determining seed viability, wherein the incubating the reaction mixture in the light chamber is carried out for 3 minutes.
Another embodiment of the present invention provides a rapid detection kit for the viability of onion seeds, wherein the kit uses changes occurring in different physiochemical parameters of onion seeds, such as enzyme activity, reactive oxygen species (ROS) concentration, free fatty acids, sugar concentration, membrane disintegration, and more, to develop markers of viable and non-viable onion seeds.
Another embodiment of the present invention provides a rapid detection kit for the viability of onion seeds, wherein by using said kit, farmers can avoid buying poor quality seeds and achieve better crop productivity. The kit can also reduce down the economic losses incurred by farmers and increase transparency in the onion seed market. This innovative idea will be highly beneficial to the Indian agricultural sector, enabling it to achieve better food security and contribute more significantly to the economy.
The following examples are provided for illustrative purpose only and do not limit the scope of the present invention.

EXAMPLES
Examples herein are for the illustration purpose and does not limit the scope of the invention in any manner.
Example:
The said method was tested for the seed varieties as – Poona Fursungi, and Bhima red. Seed samples were obtained from lots ranging from 80 to 100 kilograms. The seeds underwent a process of water washing and crushing until achieving a size of less than 400 microns. Subsequently, they were extracted with reagent A in the prescribed ratio of 1:10. In test-tubes, 1 mL of seed extract was combined with 100 µL of reagent B and exposed to light for 3 minutes. Following the incubation period, a pink-reddish coloration was observed in the test-tube containing viable seeds, while pale yellow coloration was observed in tubes containing non-viable seeds. The outcomes of this rapid method were validated against those obtained through conventional germination testing, demonstrating a consistent correlation between the two approaches.
Samples different samples were analysed and labeled as below
A: Poona fursungi lot 1
B : Poona fursungi lot 2
C: Poona fursungi lot 3
Seed Testing with rapid detection method and Conventional germination method. It is clearly visible from colour change and germination that the viable seeds showing colour change are having better germination.
Example: Spectrophotometric analysis
The method outlined herein was tested using seed varieties of Poona Fursungi and Bhima red. Two samples for each variety were tested from the different lots having variable germination. Seed samples were extracted from lots size of 80 to 100 kilograms. The seeds were subjected to water washing and crushing until achieving a size of less than 400 microns. Subsequently, they were extracted with reagent A in the prescribed ratio of 1:10. In test-tubes, 1 mL of seed extract was combined with 100 µL of reagent B and exposed to light for 3 minutes. Following the incubation period, spectrophotometric analysis was conducted at 500 nm wavelength. Seeds exhibiting a germination rate greater than 80% were expected to yield absorbance values exceeding 1.8. Thus, the efficacy of this rapid method was further confirmed by comparison with results obtained through spectrophotometric values and the germination method.
Example: Testing of onion seed for viability using the developed rapid detection kit method
A total of 4 different lots of onion seeds were tested for the seed viability using the developed rapid detection kit method.
All seed(s) were put in the wells (one seed per well) of a baseplate (101) of the kit (100). A seed crusher (102) was pressed on the seeds in the well to squeeze out the seed content. Thereafter 100ul a reagent A (103) was added to each well, plate was tapped and wait for time period for 2 minutes to 10 minutes. Furthermore, 50ul a reagent (104) B was added to each well and observed the colour change from colourless to pale yellow to pink reddish.. The number of pink-reddish wells corresponding to viable seeds were identified and counted. Finally, the seed viability was calculated.
The results obtained by said method were compared and found significantly similar with the conventional germination test. The said result was provided in table 1.
Table 1: Comparison of Rapid kit results and Germination results
Seed lot Seed Viability testing methods
Rapid kit results Germination results
1 75 % 79 %
2 43 % 44 %
3 63 % 68 %
4 32% 37%

Advantages
• Present provides rapid and reliable assessment of seed viability.
• Present invention eliminates the need for complex and time-consuming procedures.
• Present invention suitable for large seed lots.
• Present invention is provides cost effective solution to analyse seed viability.
• Present invention provides user friendly method for analysis of seed viability.
,CLAIMS:We Claim:
1. A rapid detection kit (100) for the viability of seeds, the kit (100) comprising:
i. a base plate (101) with seed wells;
ii. a seed puncher/crusher (102);
iii. a reagent A (103); and
iv. a reagent B (104);
wherein the base plate provides seed wells to place the individual seed;
wherein seed puncher presses the seed to squeeze the seed content out into the well;
wherein reagent A (103) acts as an extractant;
wherein reagent B (104) reacts with the seed extract to distinguish between viable and non-viable seeds; and
wherein the said kit provides seed viability by indicating colour change.
2. The rapid detection kit for the viability of seeds, as claimed in claim 1, wherein the kit (100) is placed in a light incubation chamber (105) for suitable time to incubate the reaction mixture.
3. A method for determining seed viability, the method comprising:
i. crushing the seeds to obtain a seed powder;
ii. contacting the seed powder with Reagent A for a time ranging from 10 minutes to 45 minutes at an ambient temperature to obtain a seed extract;
iii. filtering the seed extract to obtain a filtrate and a residue comprising suspended particles of the seeds;
iv. combining the filtrate with Reagent B to obtain a reaction mixture;
v. incubating the reaction mixture in a light chamber at light intensity in the range from 20000 lux to 30000 lux.for a time ranging from 2 minutes to 10 minutes; and
vi. observing the colour change of the reaction mixture from colourless to pale yellow or pink reddish or pink or red to determine the seed viability;
wherein colourless or pale yellow colour indicates non-viable seeds, pink reddish, pink colour to red colour indicates at least 80% viable seeds.
4. The method for determining seed viability as claimed in claim 3, wherein the seeds are washed and dried before crushing.
5. The method for determining seed viability as claimed in claim 3, wherein the seed powder is having particle size less than 400 micron.
6. The method for determining seed viability as claimed in claim 3, wherein the Reagent A is a solvents selected from water, methanol, ethanol, propanol, ethyl acetate, hexane, and combination thereof.
7. The method for determining seed viability as claimed in claim 3, wherein the ratio of seed powder to the Reagent A is 1:10.
8. The method for determining seed viability as claimed in claim 3, wherein the reagent B is azo compound or halogen compound or metal nitrates, preferably silver nitrate.
9. The method for determining seed viability as claimed in claim 3, wherein the concentration of reagent B ranges from 10mM to 200mM.
10. The method for determining seed viability as claimed in claim 3, wherein the incubating the reaction mixture in the light chamber is carried out for 3 minutes.

Dated this: May 13, 2024