cytometry are then micro-grafted to small greenhouse
grown rootstock seedlings (usually Carrizo citrange) to
expedite whole plant recovery. Use of a trifoliate rootstock
such as Carrizo allows easy identification and removal of
adventitious shoots regenerating directly from the rootstock.
Rooted plants can be transferred to any suitable
commercial potting mixture and maintained under cover at
high humidity for 2–3 weeks for acclimatization. Recovered
tetraploids should undergo molecular analysis to
determine if they are autotetraploids or allotetraploid
somatic hybrids, with both having potential value to subsequent
conventional breeding schemes. Previously,
somatic hybrids have been validated by isozymes, RAPD
or CAPS (Grosser and Gmitter 2005); more recently
unambiguous and reliable EST-SSR markers with fragments
separated on a sequencer, has been the method of
choice (Chen et al. 2008). EST-SSRs have a further
advantage of ploidy level validation in many instances, as
up to four different alleles may be identified at a single
locus in a somatic hybrid plant. Cytoplasmic genome
analysis in somatic hybrids has been done using RFLPs
(Kobayashi et al. 1991; Moreira et al. 2000), and more
recently by CAPS analysis (Guo et al. 2004b).
cytometry are then micro-grafted to small greenhouse
grown rootstock seedlings (usually Carrizo citrange) to
expedite whole plant recovery. Use of a trifoliate rootstock
such as Carrizo allows easy identification and removal of
adventitious shoots regenerating directly from the rootstock.
Rooted plants can be transferred to any suitable
commercial potting mixture and maintained under cover at
high humidity for 2–3 weeks for acclimatization. Recovered
tetraploids should undergo molecular analysis to
determine if they are autotetraploids or allotetraploid
somatic hybrids, with both having potential value to subsequent
conventional breeding schemes. Previously,
somatic hybrids have been validated by isozymes, RAPD
or CAPS (Grosser and Gmitter 2005); more recently
unambiguous and reliable EST-SSR markers with fragments
separated on a sequencer, has been the method of
choice (Chen et al. 2008). EST-SSRs have a further
advantage of ploidy level validation in many instances, as
up to four different alleles may be identified at a single
locus in a somatic hybrid plant. Cytoplasmic genome
analysis in somatic hybrids has been done using RFLPs
(Kobayashi et al. 1991; Moreira et al. 2000), and more
recently by CAPS analysis (Guo et al. 2004b).
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