The only species in which the generative nucleus has been found to
be actively involved in androgenesis is black henbane (
Hyoscyamus niger
L.). When similar looking
nuclei are formed, one or both nuclei may undergo further divisions. In some cases, the two nuclei
will fuse, producing homozygous diploid plants or callus. Since diploid callus may also arise from
somatic tissue associated with the anther, diploids produced from anther culture cannot be assumed
to be homozygous. To verify that plants produced from anther culture are haploid, chromosome
counts should be made from root tips or other meristematic somatic tissues (see Chapter 4). Because
haploids derived from diploid species are expected to be sterile or have greatly reduced fertility,
pollen staining, which is much quicker and requires less skill than chromosome counting, can also
be used to identify and eliminate potential diploids. However, pollen staining may not distinguish
between haploids and plants that have reduced fertility because they have a few extra or missing
chromosomes (i.e., aneuploids). Haploids and diploids recovered from anther culture may also be
distinguished by comparing size of cells, particularly stomatal guard cells, or through the use of
flow cytometry.