GSM2665653: exome24; Mus musculus; OTHER
For whole exome sequencing, 1–2 105 tumour cells were FACS-sorted directly into 10 lysis buffer (200 mM DTT, 100 mM TrisHCl pH 7.4, 5% SDS, 5 µg µl–1 proteinase K) and incubated at 60C for 5 h, followed by isopropanol DNA precipitation. DNA was resuspended in 0.2 mg ml–1 RNase A (Promega) and incubated for 20 min at 37C. DNA was further purified using Agencourt AMPure XP DNA beads (Beckman Coulter), washed twice in 80% ethanol and eluted in TE buffer. DNA concentration was determined using a Qubit Fluorometer (Thermo Fisher). 50 ng of gDNA from each tumour was fragmented using a Bioruptor (Diagenode) to reach an average fragment size of 300–350 bp, and fragmentation was monitored using a 2100 BioAnalyser (Agilent). Libraries for Exome capture were prepared using Kapa Library Prep Kit (KK8250) following the manufacturer´s instructions. 10 cycles of amplification. Exome capture was done by hybridization of the amplified library (200 ng/library) to the SeqCap EZ Mouse library Design Name: 110624_MM9_Exome L2R_D02_EZ_HX1 Reference #: 9999042611 (Roche Nimblegen). The purified captured DNA was then amplified by Post-LM PCR for 13 cycles.
GEO
GSM2665653
GEO Accession
GSM2665653