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‘Tomuri’ and ‘Clone A’ showed increased genetic variation based on AFLP analysis following an in vitro cycle in comparison with field-grown plants (Prado et al., 2005, 2007). Chiariotti et al. (1991) detected isoenzyme variation among plants regenerated from in vitro roots of ‘Tomuri’. Boase and Hopping (1995) obtained dodecaploid plants after axillary bud culture of hexaploid plants. Marino and Bertazza (1998) regenerated leaf-derived somaclones of A. deliciosa under different pH variations. They showed that high pH produced plants more tolerant of high pH. Protoplast culture can also produce genetic variation (protoclonal variation) in different traits of regenerated plants, e.g. cytology, morphology and resistance (reviewed by Liu et al., 2003). He et al. (1995, cited in Liu et al., 2003) reported that chromosome numbers of plants regenerated from protoplasts varied between 59 and 310, with several different ploidy levels as aneuploids, diploids, tetraploids or hexaploids. In the same work, changes in sex were reported, since a third of the plants regenerated from female A. deliciosa were male. Differences in internode length, leaf length and width and petiole length among different plants regenerated from protoplasts of A. deliciosa were also reported (Cai et al., 1992, cited in Liu et al., 2003).

Gamma-irradiation of axillary and adventitious buds was attempted for mutation breeding, but no clearly improved genotypes were obtained (Shen et al., 1990b, cited in Ferguson et al., 1996). Fraser et al. (1991) and Harvey et al. (1995) tried to achieve chromosome doubling via colchicine treatment, but very few doubled plantlets were identified. Wu et al. (2011) recovered tetraploid plants from five different genotypes of A. chinensis using colchicine treatment. In this work, small shoot buds induced from petioles were immersed in a colchicine solution (0.05 or 0.1%) for 4 h, washed with sterile water and recultured for shoot induction. Treatment with 0.05% colchicine was the optimum treatment for producing tetraploid shoots. Fruit of the autotetraploids induced from three female red-fleshed kiwifruit selections were 50–60% larger than fruit from their diploid progenitors, and this characteristic was stable over four consecutive years (Wu et al., 2012a).