Browsing by Subject "nuclei isolation"

Nuclei isolation is a method used e.g. as a part of chromatin structure research. DNA structure can be examined in its 3D form from isolated nuclei because DNA is still wrapped around the histone proteins. Examining the chromatin structure can offer information e.g. about gene expression and how it is regulated. Isolating nuclei from plant cells demand more optimization compared to animal cells because of the cell wall, chloroplasts and secondary metabolites. The presence of organellar DNA can hamper the later DNA analysis. Secondary metabolites can hinder the actual isolation process. Finding the suitable isolation protocol for species of interest may need careful optimization of different aspects. Different species can differ from each other based on the structural and biochemical characteristics and because of this the same protocol may not ensure as good results for them. Different tissue types of the same species may have also differences in their biochemical and structural characteristics. In this thesis work, three different isolation protocols were used for three plant species; Pinus sylvestris, Betula pendula and Arabidopsis thaliana. The purpose of the work was to compare the results from each nuclei isolation protocol. Optimization of isolation protocol for P. sylvestris and B. pendula would help the isolation process for later research. Thesis work was done to get guidance for this optimization. Samples handled with different protocols were different from each other based on the sample concentration (particles/mL) and the average size of isolated particles. Chloroplast contamination was tested with chloroplast specific primers with PCR. None of the samples were free from chloroplasts.