Embryogenic cells of unfertilized ovules from 24-week-old fruits of washington Navel orange were transferred to liquid MT+ 1550 mg/L glutamine + 500 mg/L ME. Protoplasts were isolated from lml habituated cell suspension culture (4-12 days into the 14 days cycle) and also from 500 mg Satsuma mandarin leavs (Citrus unshui) were transferred to CPW nutrients, Supplemented with 13% (w/v) mannitol, and a suitable combination of Celluysin and Macerozyme solution. High density of alive protoplasts were recognized by fluorescien diacetate (FDA) and using blue light microscope. Approximately equal volumes of protoplasts (105 protopast /L) in CPW were fused using PEG. After 15 minutes of dilution, the fusions were approved according to difference color of protoplasts. After 10 days, divisions of fused protoplast were stated into liquid 1BH3: 1 MT (Grosser et al, 1990). Embryogenesis and plantlet formation were followed onto MT+GA and MT+GA+ADE+ME media.
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