Isozyme Purity Testing
How to initiate a Plant Isozyme Purity Test
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About Isozyme Purity Testing
Isozyme marker loci have been available for use in quality assurance, variety identification, plant breeding, production and variety protection programs for more than four decades (43 years). Isozyme marker loci continue to be used for inbred purity, hybrid purity, inbred homozygosity determinations, inbred cleanup, integrity and hybrid cleanup. Isozyme loci are usually the markers of choice for identifying and quantifying off-types in inbred lines and selfs and off-types in single-cross hybrid populations of maize, canola, sunflowers, tomatoes, melons, etc.
Isozyme marker loci have many advantages for use in determining purity of corn or sunflower populations. First, they are not affected by the field or greenhouse environment. They are cost effective compared to other methods and the turnaround time is relatively rapid. Because at least fifteen isozyme loci can be assayed simultaneously in a single individual of maize using a single buffer system, it is possible to identify outcrosses in inbred lines and selfs and outcrosses in single-cross and multi-cross hybrid populations with a very high level of precision and at a relatively low cost. In addition, multilocus analyses provide useful information for verifying inbred and hybrid genotypes.
There are two parameters that can be varied in the laboratory when conducting purity assays. The first is the number of individuals assayed per seed lot. The second is the number of isozyme loci assayed per individual.
We have conducted many statistical analyses over time to determine the appropriate number of individuals and isozyme marker loci to assay for inbred line and/or single-cross hybrid samples of maize. The analyses routinely show that 100 random individuals are adequate in an inbred or single-cross hybrid population, as long as the number of polymorphic loci assayed is 10 or greater. Thus, BGS routinely assays a set of 11 to 15 polymorphic isozyme loci which can be assayed with a single gel buffer system.
We also have found that little is gained by assaying more than 100 random individuals with more than 11 polymorphic isozyme loci. Adding a 12th locus does not significantly improve the probability of correct classification of outcrosses (off-types) in inbred lines or single-cross hybrids or selfs in a single-cross hybrid sample, as long as 100 or more individuals are assayed.
On the other hand, if the number of plants assayed is increased to 200 or more, the number of useful (polymorphic) loci should not be decreased to fewer than nine for an optimum analysis of either inbred lines or single-cross hybrid populations. Also, if only one or two loci out of 11 to 15 loci are heterozygous in a single-cross hybrid, it is important that the other homozygous loci are assayed to optimize the opportunity to detect an outcross fertilization, which would override the interpretation of a self-fertilization based on one or a few heterozygous isozyme loci (i.e., you can not have a self if you observe an outcross genotype at any homozygous or heterozygous locus in a single individual).
To obtain further information about our molecular genetic screening, diagnostics, and GMO / GEP / Transgenic testing and Human Identity Genetics testing services and prices, contact:
Biogenetic Services, Inc. - 1321 6th Street - Brookings, South Dakota 57006