FMRP REGULATES GENOME STABILITY THROUGH DIRECT INTERACTIONS WITH R-LOOPS AND R-LOOP RESOLVASE

We performed transcriptomic analyses on pateint derived FXS cells (GM03200) compared to those derived from an unaffected individual (GM06990). EBV transformed lymphocytes derived from a normal individual (GM06990) and a Fragile X individual (GM03200) were grown in RPMI1640, supplemented with GlutaMAX, 15% heat-inactivated FBS, 100 IU/ml penicillin and 100 ug/ml streptomycin. Cells were either treated with DMSO, 0.3 micro molar aphidicolin or left untreated for 24 h before harvest. 3x106 cells were harvested for RNA-seq. RNA was extracted using the Qiagen RNeasy Plus Mini Kit. The RNA was run on an Agilent 2100 Bioanalyzer using the RNA 6000 Nano Chip to assess RNA quality and quantity. 1 µg of total RNA was used as input to the Illumina TruSeq Stranded Total RNA Library Prep Kit Ribo Zero Gold H/M/R. Library size was assessed using the DNA 1000 chip on the Bioanalyzer, and the libraries were quantified using a Qubit fluorometer. Pair-end sequencing was run on an Illumina NextSeq 500 instrument. A total of four replicates were processed for treatment/conditions out of which three were biological replicates.