GSM3703689: DCCT13.8h LZ; Mus musculus; RNA-Seq
Spleen were disrupted using a 70uM mesh strainer. Samples were then washed with FACS buffer (2% FBS and 2 mM EDTA in PBS), incubated for 30 min on ice  with antibodies and  CD43 MicroBeads (Cat# 130-049-801, Miltenyi Biotec). Red blood cells were then  lysed for 5 min at room temperature with 5mL of Gey's buffer. After a wash , samples were passed through a 70 um filter LS columns (Miltenyi) for magnetic separation and B cell enrichment. After washing,  single-cell suspension was  sorted by flow cytometry. Samples were collected in eppendorffs containing RTL Buffer Plus (Qiagen) with 1% beta-mercaptoethanol. Samples were purified with AllPrep DNA/RNA Mini and Micro Kits (Cat# 80204 and Cat# 80284, Qiagen) as per  manufacturer's instructions. Samples were prepared with Nugen cDNA kit and sequenced using Illumina HiSeq system