| description |
In this study we have sequenced the naive and antigen experienced B-cell repertoire of healthy donors of different ages as described in IJspeert et al Front. Immunol. 7:410. doi: 10.3389/fimmu.2016.00410. We have sequenced the naive repertoire from 10 healthy donors and the antigen experienced repertoire of 38 healthy donors. We have deposited both the raw sequencing results and the reads that past the filtering as described in IJspeert et al Front. Immunol. 7:410. doi: 10.3389/fimmu.2016.00410. The filtered data of the antigen selected repertoire have been devided into IGG and IGA transcripts.For the naive repertoire DNA from sorted naive B cells (CD19+ CD27-) from periperhal blood was amplified using forward VH1-6 FR1 (BIOMED-2) primers and the JH consensus reverse primer. For each healthy donor 3-6 independent PCR reaction were performed. Samples were purified by gel extraction and AMPure XP beads and sequenced using 454 GS junior sequencing with the Lib-A kit. The deposited data are untrimmed. In the analysis of IJspeert et al Front Immunol 2015 and IJspeert et al Front. Immunol. 7:410. doi: 10.3389/fimmu.2016.00410 samples were demultiplexed and filtered using CLC genomic workbench. Low quality reads (limit =0.05), reads with ambigious nucleotides (max 2), and sequences shorter than 100nt or longer than 800nt were removed. In addition the adaptor sequences were removed and sequences were trimmed 20-25 nt. From all samples both the raw data and the filtered data as described by IJspeert et al Front. Immunol. 7:410. doi: 10.3389/fimmu.2016.00410 are deposited.For analysis of the antigen experienced repertoire cDNA from the total PBMC’s was amplified using forward VH1-6 FR1 (BIOMED-2) primers and either the CgCH1 or the IGHA reverse primer. Samples were purified by gel extraction and AMPure XP beads and sequenced using 454 GS junior sequencing with the Lib-A V2 kit. Data was processed using the amplicon 1 pipeline from Roche. Samples were demultiplexed using IGGalaxy (Moorehouse et al, 2014, BMC Immunol). For all samples the raw data and the filtered data are deposited. The filtered data are divided in Cg and Ca transcripts as described in IJspeert et al Front. Immunol. 7:410. doi: 10.3389/fimmu.2016.00410. The deposited data are untrimmed. In the analysis of IJspeert et al samples were 40nt trimmed before analysis. |