Centromere plasticity and diversity: Researchers identify a novel type of centromere organization NatureComms
-type telomeres and 45S rDNA loci, respectively. Purified PCR products and plasmid DNAs were labeled with ATTO488-dUTP or ATTO550-dUTP using Fluorescent Nick Translation Labeling kits .C. japonica
were prepared from root meristems using a dropping method. Roots were pretreated in ice-cold water for 20–24 h, fixed in 3:1 fixative at RT, overnight and kept in 70% ethanol at −20 °C until use. Fixed roots were digested in an enzyme mixture , 0.7% cellulase , and 1.0% pectolyase ) in citric buffer at 37 °C for 30–40 min.
DNA replication patterns were visualized by incorporation of 5’-ethynyl-2’-deoxyuridine into the newly synthesized DNA strand using the EdU Cell Proliferation Kit . Roots ofwere incubated in a 15 µM EdU-containing Hoagland solution for 2 h at RT. They were then transferred to Hoagland solution and incubated for either 3, 6, 12 or 24 h, followed by fixation in ethanol: glacial acetic acid at RT, overnight.
leaf tissue were ground separately with liquid nitrogen and homogenized in 10 ml nuclei isolation buffer ) to isolate nuclei. Nuclei fixation was performed in 1% PFA/ nuclei isolation buffer at RT, 12 rpm for 10 min and terminated by adding 2 M glycine to a final concentration of 130 mM. The nuclei suspension was filtered through Miracloth twice and a 50 µm CellTrics filter once and centrifuged at 4 °C, 3000 ×for 10 min.
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