There are many pathways of androgenesis which leads to the formation of haploid either directly by embryogenesis or indirectly through callus formation (Fig. 6.1). Direct embryogenesis is preferred over indirect embryogenesis involving callus formation which may produce variation. There are several hundred plant species in which haploid callus or complete plantlets have been produced.
A few selected species are listed in the Table 6.1:
The pollen mother cell (P.M.C.) gives rise to pollen tetrads after meiotic division. These microspores are released in the atmosphere. The first mitotic division in the microspores gives rise to a large regenerative cell and small generative cell. The former remains quiescent while, the later divides to produce two male nuclei.
In culture although androgenesis can be induced in anthers in the tetrad stage at the binuclear pollen stage, but microspores just before or at the time of first mitosis, are most suitable for the induction of androgenesis. Examples of androgenesis are observed in datura, tobacco and barley. Androgenesis may be produced by pollen grains through mitosis producing equal or unequal nuclei.
They produce somatic embryos directly or may produce callus. Direct androgenesis is evident in Atropa belladona, Datura innoxia and Nicotiana tabacum. Indirect androgenesis is the most common method (96%) of producing haploids. This type of development (indirect androgenesis) is supposed to be result of disturbed polarity and/or complex nutrient medium.
Microspore embryogenesis has improved in recent years in case of cereals, particularly wheat, rice and barley. Direct embryogenesis (no callus phase) remains the preferred choice/route in crop improvement programmes involving hybrids.
Several factors affect the response of anther culture:
(i) The effect of genotype of donor plants i.e., genetic factor. E.g., major differences between different genotypes of wheat, rice and barley have been observed with respect to yield of callus per anther cultured and regenerant frequency of green plantlets.
(ii) Microspore Developmental Stage:
Early uninucleate, mid-uninucleate, late-uninucleate and pre-mitotic stages of microspore decide the response of anther culture, e.g., yield of callus declines when anthers bearing microspores at the stage other than mid – or late uninucleate stage are cultured.
(iii) Culture Media:
Anther culture of cereals requires modification in carbon and nitrogen components for improved anther culture, e.g. reduced concentration of ammonia nitrogen in the induction medium is effective. Sucrose, maltose and other monosaccharide as source of carbon have proved beneficial. Presence of 2, 4-D influences the response of cultures: wheat anther culture showed positive response in presence of high amount (4 mg l–1) of 2, 4-D.
(iv) Culture Methods/Conditions:
Addition of Ficoll in the culture medium increased the density and the frequency of plant formation. Aerated and proper buffered medium along with high Ficoll and sucrose amount in liquid medium make anthers float, and sustain the microspores for a longer period of time under sugar – starvation. Wei et al. (1986) indicated that sugar starvation led to induction of microspores and androgenesis.
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