In vitro expression, purification and biochemical characterization of KNL2 protein of Arabidopsis thaliana
In this diploma thesis the KNL2 homologue of Arabidopsis thaliana was characterized with biochemical methods for the first time. Recombinant Arabidopsis full-length KNL2 protein and its N- and C-terminal parts were purified and used as substrates for a kinase assay and for a DNA binding assay (EMSA). Arabidopsis KNL2 is phosphorylated by Aurora kinase 3 at N- and C-termini, and a DNA binding capability of the C-terminal part is indicated by the EMSA. The C-terminal part was used for the production of antibodies which were applied for immunostaining and Western blot experiments. The Western blot verified the specificity of the antibodies. The immunostaining showed that Arabidopsis KNL2 localizes to the chromocenters during interphase, but not during mitosis.
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