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Series GSE117800 Query DataSets for GSE117800
Status Public on Oct 01, 2018
Title Epigenetic Enhancer Marks and Transcription Factor Binding Influence Vk Gene Rearrangement in Pre-B Cells and Pro-B Cells [VDJ-seq]
Organism Mus musculus
Experiment type Other
Summary There has been no study to date that has examined genomic DNA (gDNA) and RNA Igkappa repertoires directly from the same set of cells. In this study we performed deep sequencing on Igkappa repertoire library preparations from both gDNA and RNA from the same sorted cells. This was done for both pre-B cells, where most Igkappa rearrangement takes places as well as pro-B cells where some B cells undergo early Igkappa rearrangement. In addition, we perfored gDNA repertoire analysis on iEkappa-/- pre-B cells. We find that some Vk genes display skewed DNA or RNA representiation relative to the other. We also find that pro-B cell Igkappa rearrangement is skewed to Jkappa-distal Vkappa genes. Lastly, we find that iEkappa-/- pre-B cells are not capable of rearranging to the most proximal Vkappa family genes.
Overall design gDNA (DNeasy, Qiagen) or RNA (RNeasy, Qiagen) was prepared from C57BL/6 pre-B cells, C57BL/6 pro-B cells, and iEkappa-/- pre-B cells. gDNA was sonicated to 500 to 1000 bp fragments, end repaired, A-tailed and adapter ligated. Jkappa gene biontinylated primer extended products were pulled out with streptavidin. Sequential PCRs added index, barcode and flow cell binding sequences. The last PCR utilized NEBNext Dual indexing primers. RNA was converted to cDNA with Transcriptor high-fidelity reverse transcriptase (Roche). RNase treatments removed unwanted RNA. cDNA was then adapter ligated using bridge adapters. Sequential PCRs was performed as in gDNA. Libraries were sequenced using Illumina MiSeq 2x300 read length.
Contributor(s) Kleiman E, Loguercio S, Feeney AJ
Citation(s) 30271408
Submission date Jul 27, 2018
Last update date Jan 27, 2019
Contact name Salvatore Loguercio
Organization name TSRI
Department MB
Lab Balch
Street address 10550 North Torrey Pines Rd
City La Jolla
State/province California
ZIP/Postal code 92121
Country USA
Platforms (1)
GPL16417 Illumina MiSeq (Mus musculus)
Samples (12)
GSM3309446 WT pre-B cells gDNA #1
GSM3309447 WT pre-B cells gDNA #2
GSM3309448 WT pre-B cells gDNA #3
This SubSeries is part of SuperSeries:
GSE117804 Epigenetic Enhancer Marks and Transcription Factor Binding Influence Vkappa Gene Rearrangement in Pre-B Cells and Pro-B Cells
BioProject PRJNA483208
SRA SRP155542

Download family Format
SOFT formatted family file(s) SOFTHelp
MINiML formatted family file(s) MINiMLHelp
Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE117800_RAW.tar 350.0 Kb (http)(custom) TAR (of XLSX)
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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