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| Status |
Public on Dec 16, 2017 |
| Title |
Unbiased quantification of immunoglobulin diversity at the DNA level with VDJ-seq |
| Organism |
Mus musculus |
| Experiment type |
Other
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| Summary |
For high-throughput sequencing and quantification of immunoglobulin repertoires, most methodologies utilise RNA. However, output varies enormously between recombined genes due to different promoter strengths and differential activation of lymphocyte subsets, precluding quantitation of recombinants on a per cell basis. To date, DNA-based approaches have used V gene primer cocktails, with substantial inherent biases. Here we describe VDJ-seq, which accurately quantitates immunoglobulin diversity at the DNA level in an unbiased manner. This is accomplished with a single primer extension step using biotinylated J gene primers. By addition of unique molecular identifiers (UMI) before primer extension, we reliably remove duplicate sequences and correct for sequencing and PCR errors. Furthermore, VDJ-seq captures productive and non-productive VDJ and DJ recombination events on a per cell basis. Library preparation takes 3 days, with 2 days of sequencing, and 1 day of data processing and analysis.
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| Overall design |
A range of starting material concentrations 500ng-5µg and examining different combination of DNA polymerases
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| Contributor(s) |
Chovanec P, Corcoran A |
| Citation missing |
Has this study been published? Please login to update or notify GEO. |
| Submission date |
Nov 02, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Felix Krueger |
| E-mail(s) |
fkrueger@altoslabs.com
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| Organization name |
Altos Labs
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| Department |
Bioinformatics
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| Street address |
Granta Park
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| City |
Cambridge |
| ZIP/Postal code |
CB21 6GP |
| Country |
United Kingdom |
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| Platforms (1) |
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| Samples (6)
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| Relations |
| BioProject |
PRJNA416923 |
| SRA |
SRP123508 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE106458_RAW.tar |
40.5 Mb |
(http)(custom) |
TAR (of TSV) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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