Heavy chain BCR sequence analyses of the naïve splenic BCR repertoire of WT C57BL/6 and Wiskott-Aldrich Syndrome deficient mice.

Heavy chain BCR sequence analyses of the naïve splenic BCR repertoire of WT C57BL/6 mice. Sequencing analysis was conducted using Roche 5’-454 RACE pyrosequencing platform on isolated splenic naïve CD43- B cells samples collected from pooled mice (both male and female) of both WT and Was-/- genotypes. RNA was isolated from pooled samples and were prepared using the RACE kit as well as a IgM and IgD gene specific cDNA synthesis primers. Barcoded sequences were then separated using Genious software and then filtered using IMGTHighVQuest programs to annotate individual sequence. Population statistics on clonotypic, unique sequences were then generated using the program, IgAT to determine VH family gene usage and CDR3 characteristics.