Optimisation of Immunohistochemical Detection of Cytokinins in Transgenic Arabidopsis Seedlings Expressing Activated ipt, a Cytokinin Biosynthesis Gene
| Authors | |
|---|---|
| Year of publication | 2002 |
| Type | Article in Proceedings |
| Conference | Progress in Basic, Applied and Diagnostic Histochemistry |
| MU Faculty or unit | |
| Citation | |
| Field | Morphological specializations and cytology |
| Keywords | cytokinins; immunohistochemistry; Arabidopsis |
| Description | Aseptically grown Arabidopsis seedlings were collected, fixed in PFA - GA mixture. Indirect immunohistochemistry on cryostat sections was used. Primary antibodies against zeatin-riboside and isopentenyladenine-riboside were used respectively. Goat anti-rabbit IgG conjugated with AP was used as a secondary antibody. AP activity was detected by reaction with BCIP and NBT. The best results of immunostaining were obtained by higher dilution of primary antibodies and lowering the concentration of BCIP and NBT. Arabidopsis seedlings with activated ipt showed higher the amount of reaction product in comparison with the wild type plants. Accumulation of cytokinins was established immunohistochemically during culture period in transgenic Arabidopsis seedlings, which is in accordance with the biochemical analyses. Modified immunohistochemical detection of free bases of cytokinins can be used for the semi quantitative evaluation of cytokinin content in Arabidopsis seedlings. |
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