Anther Culture of Potato and Molecular Analysis of Anther-derived Plants as Laboratory Exercises for Plant Breeding Courses

in HortTechnology

Anther culture has been one of the most successful techniques for generating haploid plants over a wide range of species. It is a reasonably simple procedure that can be accomplished successfully without sophisticated laboratory facilities; yet, the plants generated through anther culture can be used to demonstrate the application of many modern methods that have direct applicability to plant breeding. Anthers of diploid potato clones that have been selected for competence in anther culture can be cultured in a simple medium to yield androgenic embryos after 5 weeks. Plant regeneration requires an additional 3 to 4 weeks. Regenerated plants should be large enough 2 weeks after transfer to basal medium for ploidy determination by any of three methods depending on available facilities: chromosome counts in root tips; chloroplast counts in stomatal guard cells; or flow cytometry of nuclei released from in vitro plantlets. DNA can be extracted from anther-derived plantlets using a rapid extraction procedure to demonstrate segregation of PCR (polymerase chain reaction)-based markers such as RAPD (randomly amplified polymorphic DNA), RAMPs (randomly amplified microsatellite polymorphisms), or microsatellites. Microsatellite markers that were heterozygous in the anther donor can be used to verify haploidy in anther-derived plants. If an anther culture laboratory is scheduled early in a semester, such molecular analysis can be planned for late in the same semester.

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