GSM6834621: 2-4,female,control,anti-MLE,Rep2; Drosophila melanogaster; OTHER
The embryos were dechorionated in 6% bleaching solution, washed in cold 1XPBS and permeabilised using 0.05% Digitonin. pAMNase (700ng/ml) allowed to bind to antibody bound to MLE protein on chromatin. MNAse activity initiated by adding CaCl2 at 0℃ and after 30 minutes incubation allowed to stop by adding 2X RSTOP Buffer (200mM NaCl, 20mM EDTA, 4mM EGTA, 50ug/ml RNase, 40ug/ml glycogen, 10pg/ml yeast spike-in DNA). DNA fragment extracted and purified using Phenol:Chloroform extraction and 1ng of DNA used for making libraries. Library prepared using Kapa hyper prep kit using manufacturer's protocol Cut and Run sequencing, Paired end, 2X150bp illumina Hiseq