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Anther culture
Stage of development
The critical stage for successful culture in many dicot species is the first pollen mitosis.
Generally microspores are most responsive just before or at first pollen mitosis. At this
stage, tetrads of spores are released from the pollen mother cell wall and start to form the
exine (pollen outer wall). Thus the best stage generally lies between tetrad formation and
the formation of the exine.
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In some plants it is possible to correlate the stage of microspores with flower corolla length.
For example, in tobacco:
Stage
Early uninucleate
Late uninucleate
Mitotic
11
14-17
17-24
20
39
69
2 (10%)
15 (38%)
31 (45%)
0 (0%)
4 (10%)
4 (6%)
Nutritional requirements
Hormones
There appear to be two groups of plants in terms of hormone requirements.
Species requiring little or no hormones. Hormones are used sparingly because they induce
callus formation at the end of the filament, pollen, and from the anther wall instead of
direct embryogenesis from microspores. Included in this group are Nicotiana (tobacco),
Datura, Atropa, Petunia.
Species requiring hormones. Monocots such as barley and rice appear to need both auxins
and cytokinins and plantlets may regenerate from callus.
Temperature
In some cases, temperature is used to shock the microspores so that they will alter their
typical developmental pathway. For example, tobacco anthers produce more plantlets
when placed at 4 C for 5-7 days. The reported increase in response is from 21% to 58% of
anthers that give rise to embryos.
Brassica requires heat. At 25 C researchers reported that 0.5% anthers gave rise to
embryos. After culture at 35 C for 24 hours response increased to 9%.
Albino plants
In monocots such as rice and wheat, albinism can be common in plants regenerated from
anther culture. In rice, the best method found to reduce numbers of albino plants was a
cold treatment for 10-13 days at 10 to 13 C. Using cold gave 90% green and only 10%
albino embryos while a longer cold treatment gave more albinos.