Haploid plant production through anther culture in day-neutral

Haploid plant production through anther culture in day-neutral strawberry…..0.3 cm 0.5 cm 1 cma b c0.5 cm 1.5 cm 1 cmd e fg2.5 cmFig. 4. Plant regeneration by different callus ages in day-neutral strawberry cv. ‘Albion’a.1-month-old callus: small volume, soft; b. 2-month-old callus: compact, green-yellow; c: 3-montholdcallus: compact, yellow, large volume; d: 1-month-old callus after 2 months subculture, brown;e: 2-month-old callus after 2 months subculture, shoot formation; f: 3-month-old callus after 2 monthssubculture, root formation; g: plantlet on B5 medium without hormone 2 months after subculture.Arrow indicated root formationJiajun and Kai-hua (2008) indicated that the physiological condition and developmental stageof explants played an important role in regeneration. Naveed et al. (2009) reported that the highestplant regeneration frequency in wheat embryogenic-callus culture (95% by cv. ‘Chakwal -97’) wasobtained from 26 days old callus, whereas 48 days old calli showed the lowest (13%). Decreasing ofregeneration capability with increasing age of callus is a critical problem for callus maintenance andregeneration in Miscanthus x giganteus plant regeneration (Kim et al., 2010) and this finding wassimilar to our results. On the other hand, Singh et al. (2011) reported that the reduction of peroxidaseactivity can be highly correlated with increasing callus age and affected plant regeneration rate inNarigi crenulata (Roxb) culture. In strawberry, the plant regenerated from 8-week-old callus did notshow any distinct morphological variants while a significant proportion of deformed leaf shape (6 -13%) and yellow leaf (21 – 29%) was obtained among plants regenerated from 16 and 24-week-oldcalli (Nehra et al., 1992). In our own results, the 3-month-old callus was degenerated and formedroots (Fig. 3F) such as Niemirowicz-Szcytt (1990) reported.Ploidy and cytological evaluation.When evaluating cytology, the regenerants derived from anther culture had different ploidylevels: haploid, hexaploid, octoploid, and doubled-octoploid. The number of chromosomes in haploidwas 28.3 chromosomes/cell, 41.6 for hexaploid, 55.8 for octoploid, 110.6 for doubled-octoploid, and52.9 for aneuploid (Table 4 and F ig 5a; 5b; 5c). Variation in ploidy was confirmed by countingnumber of chloroplast in a stomata guard cell. Number of chloroplast per guard cell was 11.8, 17.4,20.9, 34.9, and 19.1 obtained from haploid, hexaploid, octoploid, doubled-octoploid, and aneuploidregenerants, respectively (Table 4 and Fig 5d; 5e; 5f). The ploidy ratio of regenerants via antherculture of day-neutral strawberry was 6.1% haploid (tetraploid), 3% hexaploid, 69.7 % octoploid,6.1% doubled-octoploid, and 15.2% aneuploid (Table 4).180