​Although it is acknowledged by many that the cold test for corn germination is a good predictor of field emergence, there may be confusion among what the minimum percent germination is acceptable by each company and grower. Part of the problem is range of test methods among labs, difficulty of sampling to represent the lot or even adequate sampling from a bag and characterization of delayed germinating seeds. 
 
Late emergers can have few kernels probably mostly of competition with more vigorous adjacent corn plants.  Not only do they tend to become weaker plants with smaller ears but also tend to silk when most pollen in the field has already diminished.  There have been numerous studies showing that uniform field emergence is more productive than uneven emergence.  This is true whether caused by germination problems or field problems such as compaction.
 
We are frequently asked for an opinion about the minimum standard.  I usually answer that I think the industry standard is about 85% germination but actually each company should be considering the hybrid, the test methods and their own comparisons with field studies. Uniformity of seedling emergence can only be visual factor in customer satisfaction and/or it may be a significant factor in final grain production by a hybrid, depending upon hybrid genetics and field environments of that season. Biology of maize seed is important but only one of the biological contributors to a successful and profitable season of a corn crop.

​Growing interest by corn growers in the USA to having seed tested for germination at temperatures typical of actual planting dates has resulted in several questions.  Many of these issues have been raised by seed producers for many years.  Why does some seed within a seed lot germinate and others not?  What causes some seed to not germinate under cold conditions although the warm test result shows they are alive?  What should be the minimum acceptable cold test result?
 
Hybrid corn is mostly bred to produce a large amount of grain per area of soil.  Most of that volume and weight is carbohydrate deposited in the endosperm. Genetics affecting the production and storage of carbohydrate is mostly biologically separate from the utilization of that stored carbohydrate into metabolism needed for growth of the plant.  A hybrid in which maximum grain production per plant occurs because the combination of genetics from a specific male and female parent are a match.  Those two inbreds, however, frequently differ in vulnerabilities to environments that affect germination.  The inbred that becomes the female plant in hybrid seed production becomes the genetic source of some cellular components such as the mitochondria and chloroplasts.  Mitochondria are especially significant in deriving the energy from carbohydrates to drive the metabolism needed for production of new cells during the germination process. This is probably the main biological reason that seed companies have identified certain parents as more reliable for germination.
 
Membrane systems, including those within mitochondria, are essential to cellular function.  Cellular membranes are also vulnerable to damage, especially during the rehydration after seed has be dried.  The damage is independent of temperature.  Movement of water through the pericarp into the embryo cells at the same rate in warm wet soil as in cold soil.  Sudden swelling of the membrane-bound organelles causes some breakage.  Remaining metabolism in undamaged portion repairs the damaged membranes.  This rate of repair is temperature dependent.  Repair is slower at lower temperatures.  If soil temperature is below 55°F, it is believed that very little imbibition damage to cellular membranes especially those of the mitochondria will occur.  In the field with low temperatures, the damaged seed and slow growth becomes vulnerable to invasion by microbes that can further inhibit seedling emergence.
 
Cold germination testing is intended to identify the percent of seed within a sample that have sufficient membrane damage to inhibit or delay germination and consequently not become a productive plant.  Developing reliable, repeatable test methods is a challenge resulting in multiple lab differences in results.  Some variability is due to subtle aspects of temperature of water, nature of germination media and definition of damaged seedlings usually called ‘abnormals’.  Added to the biological-environmental variables are the difficulties in adequately sampling. The initial source causing the imbibitional chilling damage vulnerability could have been disease or moisture stress in the production field, handling in the harvest, drying and bagging process.  Genetics, especially of the female parent also interact with all of these factors.  In many ways it is amazing that we produce mostly high germinating seed.

​At this time of the year in northern temperate zones, as growers are waiting for soil temperatures to be high enough for planting, one can contemplate why, no matter the position of the seed, does the root go down and the shoot go up.  This has been studied by many, including Charles Darwin in 1880.   The total mechanism is still not completely understood.
 
Geotropism is the response to gravity and phototropism is response to light.  Root’s downward growth is affected by the tissue in the root tip meristem area.  The root cap cells (outside the meristematic cells) affect the tropism.  Removal of the cap appears to disorient the growth direction.  Removal of cells on one side of the cap, causes the root to grow towards the remaining cells.  It is possible that inner cell structures such as the endoplasmic reticulum, and other cell organelles tends to become more concentrated on the lower side of the cells and thus produces metabolites on that side, resulting in cell growth in that direction. 
 
Plant hormones such as auxins and gibberellic acid are involved in plant tropism, but all of the exact mechanisms are still not understood.  It has been proposed that auxins produced in the root tip are distributed to cells behind the root tip, affecting the elongation of exterior cells and thus direction of growth.  Differences among corn varieties in root growth direction tendencies, some with more deep, narrow growth and others with more lateral growth, is evidence that genetic factors are influencing metabolism that affects root growth direction.
 
Phototropism is also affected by plant hormone production and distribution.  Specific wave lengths in natural sunlight affect the auxins involved in the differential cell elongations, ultimately resulting in growth direction towards the light source.  Exact mechanisms involving photo receptor compounds that allow specific wave lengths of light to turn on the growth cells is not clear, despite many researcher’s attempts to study specifics.
 
Tropism in plants is obvious and complex.  Surely some of those 30,000-40,000 genes in corn are involved, and mostly we can only screen for the effects of the phenomenon with selecting the best performing corn plants and admire the fact that tropism exists.

​We (some of us) think of a corn seed as an individual, as its biology is being studied.  Further, more close study can consider the individual cells and their function. Or even its important components like the mitochondria.  Considerable interest is in the DNA and how it is affecting the corn plant.  The biological interests in us concentrate on the individual as we try to understand how the corn plant functions and how to improve it.
 
This knowledge ultimately must transition into performance of multiple plants in the field.  We know that some seed will not germinate or will germinate late poorly when under cold conditions, but how does this affect overall field performance.  The late emerging plant probably will have a small, if any, ear and adjacent plants probably will not sufficiently compensate for the yield loss.  This may be significant if the season favors high yields, probably because of timely and adequate moisture, but may be insignificant in drought stressed years.
 
Corn breeders select individual plants based upon characters such as desired appearance and later based on performance in hybrids.  From those selections, specific parent and hybrid characteristics need to be considered- a mix of individual plant and field performance is used to finally decide on the commercialization of the hybrid.
 
Cold germination tests allow observations of injured or dead seed that could not recover from imbibition damage when in cold temperatures.  This information can be used to project the affect in the field.  The remaining parts of the season will determine the ideal plant density for each field.  Hybrids will vary in response to most of the complex environmental pressures of the season. Some will have root systems appropriate for the moisture allowed, some will be better silking than others, if moisture is short.  Disease and insect resistance varies among hybrids.
 
The challenge in agriculture is transitioning from knowledge of biology of individual corn plants to increasing grain production in the field.  It is obvious that this requires the effort of a lot of people with a lot of specialties.

​Germination process begins when sufficient temperature allows metabolism in the imbibed seed.  Energy transformed from the carbohydrates in the endosperm, through active mitochondria, allows the cell elongation in the root and the shoot portions of the embryo.  Geotropism and auxins guide the root downwards and the shoot upwards as the cells elongate.  The shoot apical meristem is pushed upwards as the cells away from the tip elongate forming the mesocotyl.  This continues until the tip is exposed to the red wave length of sunlight a short distance (one inch?) below the soil surface.  This light affects production of plant hormones resulting in stopping the elongation of the mesophyll cells, inducing production of the first node, with elongation of the coleoptile to push through the remaining soil.  That first node becomes the site for production of the secondary root system, as everything below that node eventually deteriorates.
 
The coleoptile, forms the ‘spike’, that emerges through the soil surface, the cells of leaves within it elongate, forcing the coleoptile to split. The first true leaves unfurl and emerge.  The first leaf is not shaped like the rest of the leaves, being shorter and growing more horizontal.  It does provide the beginning of independence from the nutrition supplied by the seed endosperm.  This first leaf will be discarded later as its energy is depleted supplying energy for the growth of the next leaves. 
 
Photosynthesis in the next few leaves provides the energy for more leaf emergence as well as the new secondary roots as they grow from that first node slightly below the soil surface.  The rate of growth of this early stage is very dependent upon temperatures.  In our greenhouse with a minimum soil of 70°F, the third leaf has emerged in about 9 days but with usual spring days in Central USA it can be 20-30 days before the third leaf is obvious.mmGenotypes vary in speed of this early growth but temperature is the major variable.
 
Success at this stage is essential for a productive crop.  Good genetics, seed quality and cooperating environment gets the corn off to a good start.

​Maize plants have from 30000 to 40000 genes according to DNA studies in which individual genes are identified according to known nucleic acids that start and stop at the ends of a gene.  Whatever is the exact number, it is assumed to be the same number for every variety of corn.  The difference between varieties are the specific codes within the gene.  This defines the genotype.   The translation of the genes often in the form of enzymes that drive the processes, interacting with the environment, results in the products that we characterize as the phenotype of the variety.
 
Phenotypes include characters critical to hybrid corn performance, such as kernel number, silking while stressed, leaf width and thickness, number of stomata, number of husk leaves, pollen production, root structure, resistance to a pathogen - all that we see when we look at a corn plant and a whole lot that we don’t see.
 
Breeding for productive and repeatable corn hybrids requires obtaining parent seed with each member of each paired chromosome having an exact genetic duplicate in the other member of the pair.  This level of homozygosity is essential to maintain duplicate future genotypes.  The process of reaching homozygosity requires selfing for several generations or short-cutting the process by use of special techniques.  Both parents, although different from each other, of a single cross hybrid must be homozygous to duplicate the hybrid genotype.
 
If the homozygous genotype or a pair of homozygous inbreds are mated, the resulting single cross plants will be identical to each other in terms of genotype.  If they are placed in a uniform environment, their phenotype will also be identical.  Some of the phenotypic characters, such as leaf shape and size, or tassel shape or branches, or color of anthers on tassel, may not affect the performance of the hybrid, or inbred, but the phenotypic character will be expressed the same if the genotypes and the environments are identical.  Some of these characters may be affected by several genes but the expression (the phenotype) will be identical if all the plants have the same genotype and identical environments.
 
Visually evaluating for identical mature plant phenotypes has long been used to evaluate whether an inbred is ‘finished’ with self-breeding before including it in hybrid production.  Observations for mature plant phenotype uniformity has also been the traditional method of evaluating genetic unity of hybrids and inbreds.  Our company (PSR) grows seedlings to the three-leaf stage to evaluate purity of inbreds and hybrids.  We have found the system is effective for all corn hybrids, regardless of genetic background.  Slight off-types due to slight genotype variation in parent seed, outcrosses due to incorrect male pollen or female plants in the seed field, self-pollinated female plants or seed mixes become evident with the private methods that we have developed over the past 32 years.  Close observation and experience, allows finding distinctions between closely related hybrid or inbreds, while also making plants available to test for specific phenotypic characters such as GMO traits.
 

Purpose of germination testing is to avoid emergence problems in the field.  All labs, private and public, attempt to have methods to accomplish this objective.  It is complicated.  A potential problem can be from a sample inadequately representing a seed lot that may include 1000 bags with 80,000 kernels per bag.  Seed sizes within a seed lot may not have the same germination quality. Not all seed lots deteriorate at the same rate, making the timing of the testing in relation to field planting can be a factor. Labs strive for consistency within the lab and many use similar methods, but subtle differences apparently influence results.
 
We participate in a referee testing in which corn seed from the same commercial bag is sent to 30-40 private and public labs for tests.  In one year’s example, the average corn warm test germination was 95.1% but the range among the 45 labs was 90.3-98.4%.  The cold tests ranged from 75-94.6% among 27 participating labs.  The germination listed on the seed bag tag could legally be 95% because seed law is based on warm test results that allow for that much variance, acknowledging all the variables involved.
 
Labs conduct a cold test in which the corn seed is planted on a paper, soil, sand or artificial soil medium, watered and kept for 7 days at 50°F before exposed to a minimum of 70°F for 4-7 days.  The principle involved is that this temperature inhibits the metabolism needed to repair damaged membranes further damaged with imbibition.  The higher initial temperatures of the warm test immediately after imbibition will allow repair of the membranes and thus normal germination and emergence.  The medium, especially concerning water-holding capacity and oxygen availability does also have an effect.  There is some confusion as to definition of saturated cold tests but it is intended to have water interfering with oxygen availability.
 
Studies that we have participated attempting to compare tests with actual field emergence generally show the cold test is the most reliable predictor of field emergence.  Over many reps, fields and years the cold test usually has the closest correlation with field emergence and that correlation value is often about 70%. Obviously other field factors also influence emergence.
 
Our company (PSR) performs many tests for companies and growers.  400 seeds are planted in an artificial soil mix prepared for us.  It includes a surfactant to allow even distribution of water and yet enough aeration for seeds to respire.  Cold test samples are planted, watered and placed in a 50°F cold room for 7 days.  Samples then are placed on a greenhouse bench heated with a minimum of 70°F until the third leaf is visible for most seedlings.  All labs struggle with classification of the few seed that emerge late and perhaps show some injury in the first leaves.  Our policy is to characterize those seed that only show the ‘spike’ when most are at 3 leaves as not germinating.  We assume that even if these individuals do germinate late in the field, they will be non-performing plants because of competition with adjacent plants.
 
PSR warm tests tend to give lower results than many labs, but not the lowest among referee labs, probably because of our characterization of the late emergers and especially with marginal quality lots.  Our cold tests tend to be about average of the wide range of lab results.

​Aerobic respiration is the process in which oxygen molecules are utilized in breaking loose the stored chemical energy of carbohydrates into the biological usable energy chemical of ATP (adenosine triphosphate).  This series of chemical reactions is called the citric acid cycle or Krebs cycle.  Oxygen is absorbed in the cycle ultimately ending in the bi-products of CO2 and H2O.
 
This activity occurs in the mitochondria of cells on their membranes.  This process is common to all aerobic organisms, like us and corn seeds.  It is ultimate process in which we and corn seeds obtain the energy for all biological functions.  Maize seed stored at low internal moisture levels have low respiration rates but, upon imbibition and adequate heat, the Krebs cycle rate dramatically increases.  This can be evaluated by measuring O2 uptake and CO2 discharge in controlled environments.
 
A research paper published in 1967 (Plant Physiol. 42, 1071-1076) compared the O2 uptake levels during first hours after imbibition and first days of seed germination. After comparing seed at different levels of damage from storage conditions, the research indicated that the initial oxygen uptake was related to eventual metabolism of the germinating seed 3-5 days later.
 
I interpret this and later studies to indicate that seed germination success is dependent upon mitochondrial membrane integrity, especially during imbibition.  Individual cells include large numbers of mitochondria, and the embryo includes large numbers of cells, it seems reasonable that those individual seed that germinate slowly have some percentage of damaged mitochondria.  Respiration rates on intact mitochondrial membranes are affected by heat energy, allowing the ATP to assist in membrane repair. Corn seed respiration rates increase with temperatures above 50°F as reflected in warm and cold germination tests.
 
Imbibition occurs at any temperature, but respiration activity is dependent upon mitochondrial membrane integrity and sufficient oxygen and heat energy to provide the ATP for all other cellular functions involved in seed germination.