Journal
EUPHYTICA
Volume 173, Issue 2, Pages 235-242Publisher
SPRINGER
DOI: 10.1007/s10681-009-0098-y
Keywords
Autotetraploid; Cell suspension; Citrus reticulata; Colchicine; Enzymatic maceration; Flow cytometry
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An efficient protocol for colchicine mediated production of in vitro autotetetraploids from Ponkan mandarin using cell suspension cultures is described. Cells were treated with 1 g l(-1) colchicine for 4 or 8 days before transfer into solid EME medium supplemented with 5% maltose. Colchicine treated cells were placed in medium with or without an overlay of 1:2 medium-mixture of liquid 0.6 M BH3 medium and 0.15 M EME + maltose liquid media. It was observed that modifying the immediate cell environment by addition of the liquid overlay played a positive role in cell differentiation and subsequent plant regeneration. Ploidy levels were determined with a flow cytometer and confirmed by chromosome staining using the enzymatic maceration method. A large number of non-chimeric autotetraploids were generated using this method. Such plants have great value in a breeding program for the development of seedless triploid citrus, as very few available tetraploid breeding parents are easy to peel.
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