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Fig. 7. Changes in DNA loops in lamin-depleted cells. The structure of DNA halos from lamin-depleted cells was analysed (A-C). Cells were depleted of lamin A/C or B1 using RNAi vectors and co-transfection with a vector expressing an appropriate fluorescent-lamin A/C or B1, as reporter that marks the nuclear periphery. After 36 hours, nucleoids were prepared and samples containing ethidium bromide (A) used to define halo structure and the corresponding DNA loop length (C). Time-lapse epi-fluorescence microscopy was also used to monitor the expansion of DNA halos, using 15 second time frames over 2 minutes; two frames from typical time-lapse series at 30 and 90 seconds are shown (B). DNA halos prepared from lamin A/C depleted cells expanded to give uniform halos of DNA loops of 19.1±3.9 µm (n=58), which is equivalent to an average DNA loop, measured from the nuclear rim, of 112 kbp; this was indistinguishable from the DNA loops of untransfected controls in the same preparation (A). The average halo from lamn-B1-depleted cells was 24.0±5.5 µm (n=64), equivalent to 141 kbp. White arrows highlight abnormal structures seen in lamin-B1-depleted cells. The size of scale bars is given on individual images. Scale bars: 20 µm.
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