Viability

A viable seed is one, which is capable of germinating under the proper circumstances. Such a viable seed may or may not be readily or immediately germinable. Dormant viable seeds may require lengthy specific treatments before they become immediately germinable.

Topographical tetrazolium or TZ test and embryo excision tests are two test methods of seed viability. Topographical tetrazolium or TZ test is very useful for rapidly obtaining an indication of germination potential and viability of samples and is in extensive use. Embryo excision tests is to determine quickly the viability of tree seeds which normally germinate slowly, or show dormancy under the prescribed methods to such an extent that a complete germination test requires more than 60 days. The test is not valid for previously germinated seeds and must not be applied to submitted samples, which contain any dry germinated seed.

TZ Test

In this biochemical test, living cells are made visible by reduction of an indicator dye. The indicator used in the TZ test is a colourless solution of a tetrazolium salt imbibed by the seed. Within the seed tissues, it interferes with the reduction processes of living cells and accepts hydrogen from the hydrogen from the hydrogenases. By hydrogenation of the 2,3,5-triphenyl tetrazolium chloride, a red, stable and non-diffusable substance, triphenyl formazan is produced in living cells.

Testing sample

A representative sample of fifty or one hundred seeds is usually sufficient for most practical tetrazolium tests.

Staining

The prepared seed should be placed in suitable container and covered with the testing solution, and keeping it in a dark, warm place (40⁰C). When observations indicate that a sample has stained sufficiently, the TZ solution should be discarded and the seed sample covered immediately with water. Refrigeration should be used to preserve samples that are not to be interpreted within a few hours. Tests can usually be kept readable for at least three days at 10⁰C.

Evaluation of samples

The sample is ready for evaluation when it is stained. The evaluation methods for different groups as below:

• Group A- Maize, Sorghum, Small grains

Magnification of about 5-7X is desirable.

A germinable seed

The embryo structures are well-developed, non-fractured and of a normal cherry red colour; vascular tissue is deeply stained; scutellum in corn stippled in appearance, with dark red dots against a lighter red background, root-shot axis obviously swollen and plumule curled away form scutellum, when test is allowed to stand several hours.

1. The embryo contains no more than the maximum listed for one or more of the following types of deterioration.

1. Necroses within the upper or lower ends of the scutellum. No more than one-third of the scutellum should be unstained at either end.

2. Radicle unstained (Sorghum is an exception. Sorghum seeds do not have seminal roots, so it is essential that radicle be normally stained)

3. Shallow layer of white, blurred tissue over the cut surface of embryo structures.

A non-germinable seed

1. Shoot largely unstained.

2. Scutellar node unstained.

3. Major areas of coleoptile unstained.

4. Central area of scutellum unstained.

5. Insect, rodent, mechanical or other injures causing one or more essential structures to be non-functional.

6. Deep-seated layer of white, blurred tissues along cut surfaces.

7. Freezing injury.

• Group B-Large-seeded grasses

Magnification of 7-10 X is desirable.

A germinable seed

1. The embryo structures are well developed, non-fractured and of a normal red colour.

2. The embryo contains no more than the maximum listed for one or more of the following types of deterioration.

1. No more than one-third of the scutellum should be un stained at either end.

2. Radicle unstained.

A non-germinable seed

1. The complete embryo, or a major portion of it, is non-stained or of distinctly abnormal colour or texture:

2. Embryo with completely non-stained root meristem and/or plumules.

3. Embryo with more than one-third of the scutellum unstained at either end or the mid-section.

4. Embryo structures with brownish-red, soft, poorly outlined boundaries.

5. Embryo immature or absent.

• Group C-Small-seeded grasses

Magnification of 10-15 X is necessary.

A germinable seed

1. Embryo structures are well-developed, non-fractured, and of a normal red colour.

2. The embryo contains no more than the maximum listed for the following evidences of deterioration:

(a) lightly stained but with distinctly outlined embryos.

(b) basal portions of radicle and scutellum unstained, provided the embryonic axis and remainder of scutellum are well-stained.

A non-germinable seed

1. Embryo missing, immature or unstained.

2. Embryo axis unstained but scutellum stained.

3. Upper or lower half of embryo unstained.

4. Embryo dark ,dulls or pales red, with endosperm of a yellowish or greenish colour.

5. Embryo deep red with blurred outline.

• Group D- Legumes

Magnification of 7 X desirable for the small-seeded species but not necessary for larger seeds.

A germinable seed

1. The embryo is well-developed, non-fractured, and of a normal red colour and condition

2. The embryo contains no more than the maximum listed for one or more of the following

1. Small, shallow, unstained or intensively stained areas on outer sufaces of cotyledons.

2. One cotyledon completely fractured at point of attachment, or complete transverse fracturing of both cotyledons with no more than one-half of the cotyledon tissue non-functional.

3. Unstained areas near the embryonic axis attachment on either cotyledon, which does not involve vascular tissues of at least one cotyledon.

4. Shallow non-stained areas on hypocotyls.

5. In peanuts, radicle tip white or deep red up to three-fourths of the triangular section of the longitudinally bisected stele.

6. Necroses or dark, watery, red areas on the outer surface of the hypocotyls, especially frequent in snap beans and soybeans, provided they have not extended deeply into the stele.

7. Embryo with yellow or a faint pink cast, if firm in texture, and giving evidence of delayed absorption of tetrazolium.

8. Less than one-half of cotyledon tissues unstained or non-functional.

A non-germinable seed

1. The complete embryo, or a major portion of it, is not stained and is of dull appearance and flaccid, or is of distinctly abnormal colour or texture.

2. Embryo with deep-seated deterioration of cotyledon tissues when slight pressure is applied.

3. Embryo with deep-seated deterioration of cotyledon tissues that extends to inner flat surfaces.

4. Embryo with both cotyledons functionally severed from embryonic axis by fractures or deteriorated tissues, or by transverse fractures or deteriorated areas that cause more than one-half of the total cotyledon tissues to be non-functional.

5. Embryo with extensive surface necroses involving vascular tissues, or extensive mottling of brownish or bluish red and white staining patterns.

6. Embryo with deteriorated areas on hypocotyls that involve more than one-half of the diameter of the stele.

7. Embryo with deterioration of radicle that extends, upward and beyond the tapering, or angular cell division area, of the stele.

8. Embryo with epicotyl, or both plumules, made non-functional by fractures.

9. Embryo with necroses of plumules, especially frequent in snap beans, that cause more than one-half of the plumule surfaces to be non functional. In borderline cases, the pair of plumules should be broken loose for observation on both sides.

• Group E-Dicotyledonous seeds other than legumes

This is a heterogeneous group of species.

1. The embryo structures are well-developed, intact and of normal red colour.

2. Embryo contains no more than the maximum listed for one or more of the following:

1. Small necroses on cotyledons in areas other than at juncture of embryonic axis and cotyledons.

2. Small necroses at the extreme tip of the radicle.

A non-germinable

1. Embryo completely or mostly unstained.

2. More than extreme tip of radicle unstained.

3. More than one-half of cotyledon tissue unstained or made non-functional by a fracture.

4. Deep-seated necrosis at cotyledon and embryonic axis juncture, or on radicle.

5. Purplish-red or greyish-red stain.

6. Fractured radicle.

7. Poorly developed immature seed.

Embryo excision method

The object of the excised embryo test is to determine quickly the viability of tree seeds, which normally germinate slowly, or show dormancy under the prescribed methods to such an extent that a complete germination test requires more than 60 days. The test is not valid for previously germinated seeds and must not applied to submitted samples which contain any dry germinated seed.

• Method

The test is performed on four replicates of fifty seeds drawn at random from the pure seed fraction of a purity test. The seeds are soaked for one to four days in slowly running water or in standing water at a temperature below 25 0 C, or in standing water at room temperature, with at least two changes of water per day. The embryos are excised from the soaked seeds under moderately sterile conditions in a clean drought-proof room e.g. below a sheet of glass fixed about 200 mm above the working surface.

• Incubation

The excised embryos should be placed on top of filter paper and kept under normal conditions of light and moisture at a constant temperature of 20 C to 25 C for up to fourteen days. The seeds or fruits should be soaked for fifteen minutes in a 5% sodium hypochlorite solution, and washed well water before excision.

• Evaluation

The embryos should be examined daily, and differentiation between viable and non-viable embryos can be made, up to a maximum of fourteen days.

The following categories

1. Germinating embryos

2. Embryos with one or more cotyledons exhibiting growth or greening.

3. Embryos remaining firm, slightly enlarged and either white or yellow according to species.

4. Embryos of conifers that exhibit curvature of the hypocotyl.

• Non-viable

1. Embryos, which rapidly develop server, mold, deteriorate and decay.

2. Degenerated embryos

3. Embryos exhibiting extreme brown or black discolouration and off-grey colour or white watery appearance.

4. Dead or embryoless seeds detected during preparation and seeds with deformed embryos.

The seed viability in a number of tree spp has been successfully deduced through X-ray photography.

Vigour

Vigour is the sum total of all seed attributes which favour rapid and uniform stand establishment in the field.

• Criteria

1. Reproducibility of results, which could be objectively assessed, clear and meaningfully interpreted.

2. The correlation with the field stand, that is, seedling emergence.

3. It should be based on sound principles.

4. It should be simple and cheap to perform on a large number of samples in rapid succession.

The ultimate proof of any Vigour test is its reliability in predicting field stands under a variety of field conditions.

• Kinds of Vigour Tests

1. Direct Tests: it simultaneously evaluates all the factors affecting seed Vigour.

2. Indirect Tests: these tests measure certain physiological attributes of seeds. These test have the advantage that the variables can be precisely controlled, allowing reproducibility of results. They also allow direct vigour comparisons over a wide geographic area.

• Direct method

Brick gravel test

The first vigour test to gain widespread acceptance was the brick gravel test. In vigour tests for small grains, about 30 mm layer of moist gravel is placed above the seed. This layer impedes the emergence of weak, partially diseased. This layer impedes the emergence of weak, partially diseased, corkscrew-type seedlings, and other seedlings with injured coleoptile tips. The inter-wedging of the brick gravel places a stress upon the emergence of elongating shoots. The seedlings that emerge through the layer of brick gravel are considered vigorous.

Paper piercing test

This method utilizes regular testing sand plus a specially selected type of paper disk through which seedlings penetrate in order to be considered vigorous. The test as used for cereal crops involves placing seed on top of approximately 1.25 cm of moist sand, covering of seeds with a specially selected type of paper disk and then covering of paper with about 3 cms of moist sand. Tests are held at 20 0C for eighty days.

Accelerated ageing

In this test, prior to placing the seeds for germination tests, seeds are aged at high temperatures ranging from 40 to 450C and 100% relative humidity, for periods ranging up to seven days,

The cold test for corn

It provides information not obtainable from the standard germination test. Consideration of both cold test responses and standard germination responses provides the grower, dealer or farmer with a clearer insight into the quality of corn seed lots.

Dry weight of seedlings

This method is particularly adapted to grasses. Seedlings are grown in flats in a greenhouse for a period of five or six weeks. Seedlings are cut off at ground level with a razor blade, dried at 1000C for twenty-four hours and weighed. If seed germination is low, determinations on a weight per seedling basis may be more meaningful than on per hundred seed basis.

Speed of germination

The seed lots which produce the largest number of germinated seeds at the preliminary count will produce the fastest growing seedlings and the fastest stand establishment.

Seedling length measurements

The tests are placed in germinators at a 45 0 angle. After an appropriate period (five days at 250C for corn, six days at 20 0C for wheat), the length of roots or shoots is measured with a ruler. The average length of seedlings per sample is calculated. Six replications of fifteen to twenty-five seeds each is desirable.

Exhaustion test

The exhaustion test involves germination and seedling development in complete darkness, and with carefully regulated amounts of water supply.

Conductivity test

This test is more commonly being used for determining the seed vigour in crops such as, wrinkled seeded garden peas. This is based on the principle that seeds which are loosing vigour release materials, such as sugars, or other electrolytes, in solution into the soil which may increase the activity of soil fungi, which inturn, may interfere with the development of the seedling growth, especially under cold and wet conditions.

The weights and conductivity readings for the test replicates of a sample are averaged, and the conductivity per gram of seed weight is calculated. The samples, which give high reading, have been found to be correlated with low field emergence. If the very high values are obtained the seeds may not be suitable for sowing.

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