The effect of polyamines on the centromeric region of the mitotic chromosome

Current models of mitotic chromosome organization have focused mainly on the role that different the proteins have. However, recent studies have found that polyamines (PA) also have an effect on the condensation of the mitotic chromosome. It was reported that without PA the mitotic chromosome swells and gains volume. When PA is reintroduced the initial volume is recovered. In this thesis, the effect of PA on the condensation of chromosomes was further investigated. First, the effect that changing the PA content of the buffer has on the entire mitotic chromosome was investigated. It was observed that when a chromosome is moved from a buffer with PA to a buffer without PA the length of the chromosome is increased. However, the reverse action was not observed. Next, the effect that PA has on protein rich regions of the mitotic chromosome was investigated. To this end, staining of a variety of proteins with immunofluorescence (IF) staining was tried. The proteins of interest were,...Show moreCurrent models of mitotic chromosome organization have focused mainly on the role that different the proteins have. However, recent studies have found that polyamines (PA) also have an effect on the condensation of the mitotic chromosome. It was reported that without PA the mitotic chromosome swells and gains volume. When PA is reintroduced the initial volume is recovered. In this thesis, the effect of PA on the condensation of chromosomes was further investigated. First, the effect that changing the PA content of the buffer has on the entire mitotic chromosome was investigated. It was observed that when a chromosome is moved from a buffer with PA to a buffer without PA the length of the chromosome is increased. However, the reverse action was not observed. Next, the effect that PA has on protein rich regions of the mitotic chromosome was investigated. To this end, staining of a variety of proteins with immunofluorescence (IF) staining was tried. The proteins of interest were, centromere protein A (CENPA), NCAPH1, NCAPH2 and Topoisomerase IIa. Whilst early test of CENPA staining were successful this result was not reproducible when new antibodies were ordered. Furthermore, the staining of the other proteins was also unsuccessful. All of the unsuccessful IF staining showed a lack of specificity since most of the chromosomes were stained instead of the specific regions were the proteins are known to aggregate. Since no successful staining of proteins could be performed, it was not possible to measure the effects that polyamines have on the regions were CENPA, NCAPH1, NCAPH2 and Topoisomerase IIa bind.Show less

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The effect of polyamines on the centromeric region of the mitotic chromosome