Allotetraploid hybrids produced by protoplast fusion for seedless triploid Citrus breeding

Protoplast fusion has been used to produce novel allotetraploid hybrids for use as parents in crosses with diploids to produce easy-peel, seedless, triploid citrus cultivars. Forty three plants were produced from fusions between protoplasts of 'Encore' mandarin and 'Valencia' sweet orange, and of 'Encore' and 'Caffin' clementine mandarin in which protoplasts of each of the donors were isolated from embryogenic callus. Forty plants were confirmed as tetraploid by flow cytometry and chromosome counting. Three ( less than 10% of all regenerants) were non-hybrid diploids. DNA fingerprinting confirmed that both parents contributed DNA to the hybrids, which were therefore allotetraploid. The hybrids showed morphological features typical of other tetraploid citrus, including broad, thick, dark green leaves. However, they grew much faster than colchicine-induced autotetraploids. The allotetraploid hybrids will be used in crosses with the diploid cultivar 'Encore' mandarin and other clementine-type mandarins in an effort to generate seedless, triploid, easy-peel cultivars with 'Encore-type' flavor.