Analysis of polyclonal C57BL/6 repertoires. In 2 independent analyses, C57BL/6 splenocytes were sorted into CD4+GFP-Foxp3- and CD4+GFP-Foxp3+ populations and the Vβ8.2 TCR repertoire analyzed. Frequency of total (A) and unique (B) sequences acquired for each analysis without or with filtering sequences at q = 30. For each unique sequence acquired, sequences present at lower frequency with a single nt mismatch were tabulated. For the 20 most frequent sequences in each cohort, the total number of single nt mismatch sequences present at less than the indicated frequency (abscissa) relative to each corresponding high frequency index sequence were tallied. The total number of these presumed erroneous sequences for the Foxp3- (C) and Foxp3+ (D) populations either analyzed without filtering or filtered at a q = 30 are plotted (ordinate). Results demonstrate a decreased number of presumed erroneous sequences after applying a q = 30 filter. (E) For each unique sequence, the total number of other unique sequences present at a lower frequency and with a single nt mismatch was tallied. The number of these single mismatch sequences was summed for all sequences within each cohort with or without q = 30 filtering. (F) ACE values were calculated as estimates of total repertoire diversity in populations either with or without q = 30 filtering.
Nguyen et al. BMC Genomics 2011 12:106 doi:10.1186/1471-2164-12-106