Anther culture of maize and the visualization of embryogenic microspores by fluorescent microscopy

Abstract

Three maize genotypes previously shown in the literature to respond to anther culture were tested under various conditions. Studies indicated that embryogenic response ranged from 0 to 100 embryos per 1,000 anthers plated and was significantly lower without cold pretreatment of the anthers. Culture in liquid media tended to produce more embryos than in semi-solid as did the addition of activated charcoal to either liquid or solid culture media. Most results were confounded by plant-to-plant variation which tended to obscure significant differences. In one study, germination rate of androgenetic embryos averaged about 20%, but only 26% of those embryos that germinated completed their reproductive cycle and formed seed albeit through sibpollination since plants could not be selfed. Chromosome counts using root tip squashes indicated that regenerated plants were either haploid or diploid but plants scored as non-diploid yielded as much seed as scored diploids. This suggests that progeny can be recovered even from putative haploids, presumably as a result of "sectoring" in the developing ear. A DNA-specific fluorescent dye was used to visualize the presence of putative embryogenic microspores (PEMs) during the culture period. PEM counts were a function of time in culture and were apparently greater than the number of embryos obtained for a given treatment. The data indicate that, as previously reported for other species, both induction and survival phases also exist in maize anther culture.

Doubled Haploid Production in Capsicum Annuum L. Using Anther Culture : A Review

“…Flower bud whose corolla is slightly larger than sepals contain microspores having partial anthocynin colouration are said to be of later stages (Mityko et al, 1995;Novak, 1974;Morrison et al, 1986a). Dolcet-Sanjuan et al (1997) confirmed the development stage of microspores (10% binucleate) using the method by Pace et al (1987), by staining with mithramycin A. DAPI staining was also done for best stage determination (Coleman and Goff, 1985). Lantos et al (2009) has experimented with collected buds of late uninucleate and early binucleate stages of microspore development for effective embryogenesis to occur in C. annuum.…”

Isolated wheat microspore culture

“…Microspore viability was measured after isolation using the fluorescent stain, FDA (fluorescein diacetate - Pace et al 1987) and recorded as the number of viable and nonviable cells (FDA counts) and converted to a percentage for analysis. To determine the isolation procedure which gave the highest viability, four isolation procedures (blending, floating, macerating and stirring), and either pre-isolation mannitol conditioning or no mannitol conditioning (4 x 2 ~ 8 isolation treatments) were evaluated.…”

Cytological aspects of in vitro androgenesis in wheat (Triticum aestivum L.) using fluorescent microscopy

“…Such an exact differentiation was not always possible in the acetocarmine stained samples (Henry et al 1984). The application of DNA-specific fluorescent dyes in different fields of pollen biology (Coleman and Golf 1985;Hough et al 1985;Vergne et al 1987) and biotechnology (Kyo and Harada 1986;Pace et al 1987) is very advantageous.…”