Project description:DNA topoisomerase II (topo II) catalyzes a strand passage reaction in that one duplex is passed through a transient brake in another. Completion of late stages of neuronal development depends on the presence of active isoform (topo IIbeta). We identified topo IIbeta action sites on 7 selected genomic regions (about 79 Mb in total), each containing at least one gene that is controlled by topo IIbeta in neuronal differentiation. We used a novel method, etoposide-mediated topoisomerase immunoprecipitation (eTIP), followed by identification of precipitated DNA fragments on genomic tiling arrays. These DNA fragments were first fractionated by concentrated salt prior to the array analysis. The 0.5 M NaCl-released fraction was analyzed as P2 and the salt-resistant fraction wad analyzed as P1.
Project description:DNA topoisomerase II (topo II) catalyzes a strand passage reaction in that one duplex is passed through a transient brake in another. Completion of late stages of neuronal development depends on the presence of active isoform (topo IIbeta). We identified topo IIbeta action sites on 7 selected genomic regions (about 79 Mb in total), each containing at least one gene that is controlled by topo IIbeta in neuronal differentiation. We used a novel method, etoposide-mediated topoisomerase immunoprecipitation (eTIP), followed by identification of precipitated DNA fragments on genomic tiling arrays. These DNA fragments were first fractionated by concentrated salt prior to the array analysis. The 0.5 M NaCl-released fraction was analyzed as P2 and the salt-resistant fraction wad analyzed as P1. The genomic DNA fractions from the immunoprecipitate (P1 or P2) were co-hybridized with a reference DNA isolated from the input of immunoprecipitation.
Project description:Expression analysis was performed with in vitro cultured cerebellar granule neurons (CGNs) isolated from rat brain. The CGNs were culture for four weeks. Each sample was collected after interval of seven days. No treatment was given to any cultured neurons at any time point. The purpose of the experiment was to identify the genes differentially expressed during the senescence of CGNs. The experiment is useful in revealing the senescence associated genetic markers in neurons.
