دورية أكاديمية
Enriching and Characterizing T Cell Repertoires from 3' Barcoded Single-Cell Whole Transcriptome Amplification Products.
| العنوان: | Enriching and Characterizing T Cell Repertoires from 3' Barcoded Single-Cell Whole Transcriptome Amplification Products. |
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| المؤلفون: | Jivanjee T; Institute for Medical Engineering & Science and Department of Chemistry, Massachusetts Institute of Technology, Cambridge, MA, USA.; Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA, USA.; Ragon Institute of MGH, MIT, and Harvard, Cambridge, MA, USA.; Broad Institute of MIT and Harvard, Cambridge, MA, Massachusetts Institute of Technology, Cambridge, MA, USA., Ibrahim S; Institute for Medical Engineering & Science and Department of Chemistry, Massachusetts Institute of Technology, Cambridge, MA, USA.; Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA, USA.; Ragon Institute of MGH, MIT, and Harvard, Cambridge, MA, USA.; Broad Institute of MIT and Harvard, Cambridge, MA, Massachusetts Institute of Technology, Cambridge, MA, USA., Nyquist SK; Institute for Medical Engineering & Science and Department of Chemistry, Massachusetts Institute of Technology, Cambridge, MA, USA.; Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA, USA.; Ragon Institute of MGH, MIT, and Harvard, Cambridge, MA, USA.; Broad Institute of MIT and Harvard, Cambridge, MA, Massachusetts Institute of Technology, Cambridge, MA, USA.; Program in Computational and Systems Biology, Massachusetts Institute of Technology, Cambridge, MA, USA.; Computer Science and Artificial Intelligence Laboratory, Massachusetts Institute of Technology, Cambridge, MA, USA., Gatter GJ; Institute for Medical Engineering & Science and Department of Chemistry, Massachusetts Institute of Technology, Cambridge, MA, USA.; Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA, USA.; Ragon Institute of MGH, MIT, and Harvard, Cambridge, MA, USA.; Broad Institute of MIT and Harvard, Cambridge, MA, Massachusetts Institute of Technology, Cambridge, MA, USA., Bromley JD; Institute for Medical Engineering & Science and Department of Chemistry, Massachusetts Institute of Technology, Cambridge, MA, USA.; Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA, USA.; Ragon Institute of MGH, MIT, and Harvard, Cambridge, MA, USA.; Broad Institute of MIT and Harvard, Cambridge, MA, Massachusetts Institute of Technology, Cambridge, MA, USA.; Microbiology Graduate Program, Massachusetts Institute of Technology, Cambridge, MA, USA., Jaiswal S; Department of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, MA, USA., Berger B; Broad Institute of MIT and Harvard, Cambridge, MA, Massachusetts Institute of Technology, Cambridge, MA, USA.; Program in Computational and Systems Biology, Massachusetts Institute of Technology, Cambridge, MA, USA., Behar SM; Department of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, MA, USA., Love JC; Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA, USA. clove@mit.edu.; Ragon Institute of MGH, MIT, and Harvard, Cambridge, MA, USA. clove@mit.edu.; Broad Institute of MIT and Harvard, Cambridge, MA, Massachusetts Institute of Technology, Cambridge, MA, USA. clove@mit.edu.; Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA. clove@mit.edu., Shalek AK; Institute for Medical Engineering & Science and Department of Chemistry, Massachusetts Institute of Technology, Cambridge, MA, USA. shalek@mit.edu.; Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA, USA. shalek@mit.edu.; Ragon Institute of MGH, MIT, and Harvard, Cambridge, MA, USA. shalek@mit.edu.; Broad Institute of MIT and Harvard, Cambridge, MA, Massachusetts Institute of Technology, Cambridge, MA, USA. shalek@mit.edu.; Program in Computational and Systems Biology, Massachusetts Institute of Technology, Cambridge, MA, USA. shalek@mit.edu. |
| المصدر: | Methods in molecular biology (Clifton, N.J.) [Methods Mol Biol] 2022; Vol. 2574, pp. 159-182. |
| نوع المنشور: | Journal Article |
| اللغة: | English |
| بيانات الدورية: | Publisher: Humana Press Country of Publication: United States NLM ID: 9214969 Publication Model: Print Cited Medium: Internet ISSN: 1940-6029 (Electronic) Linking ISSN: 10643745 NLM ISO Abbreviation: Methods Mol Biol Subsets: MEDLINE |
| أسماء مطبوعة: | Publication: Totowa, NJ : Humana Press Original Publication: Clifton, N.J. : Humana Press, |
| مواضيع طبية MeSH: | T-Lymphocytes* , Transcriptome*, Receptors, Antigen, T-Cell/genetics ; Sequence Analysis, RNA/methods ; Single-Cell Analysis/methods |
| مستخلص: | Antigen-specific T cells play an essential role in immunoregulation and many diseases such as cancer. Characterizing the T cell receptor (TCR) sequences that encode T cell specificity is critical for elucidating the antigenic determinants of immunological diseases and designing therapeutic remedies. However, methods of obtaining single-cell TCR sequencing data are labor and cost intensive, typically requiring both cell sorting and full-length single-cell RNA-sequencing (scRNA-seq). New high-throughput 3' cell-barcoding scRNA-seq methods can simplify and scale this process; however, they do not routinely capture TCR sequences during library preparation and sequencing. While 5' cell-barcoding scRNA-seq methods can be used to examine TCR repertoire at single-cell resolution, doing so requires specialized reagents which cannot be applied to samples previously processed using 3' cell-barcoding methods.Here, we outline a method for sequencing TCRα and TCRβ transcripts from samples already processed using 3' cell-barcoding scRNA-seq platforms, ensuring TCR recovery at a single-cell resolution. In short, a fraction of the 3' barcoded whole transcriptome amplification (WTA) product typically used to generate a massively parallel 3' scRNA-seq library is enriched for TCR transcripts using biotinylated probes and further amplified using the same universal primer sequence from WTA. Primer extension using TCR V-region primers and targeted PCR amplification using a second universal primer result in a 3' barcoded single-cell CDR3-enriched library that can be sequenced with custom sequencing primers. Coupled with 3' scRNA-seq of the same WTA, this method enables simultaneous analysis of single-cell transcriptomes and TCR sequences which can help interpret inherent heterogeneity among antigen-specific T cells and salient disease biology. The method presented here can also be adapted readily to enrich and sequence other transcripts of interest from both 3' and 5' barcoded scRNA-seq WTA libraries. (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.) |
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| معلومات مُعتمدة: | R35 GM141861 United States GM NIGMS NIH HHS; T32 GM087237 United States GM NIGMS NIH HHS |
| فهرسة مساهمة: | Keywords: Gene expression; Seq-Well S3; Single-cell RNA-sequencing; Single-cell TCR sequencing; T cell receptor repertoire profiling; Targeted enrichment; scRNA-seq |
| المشرفين على المادة: | 0 (Receptors, Antigen, T-Cell) |
| تواريخ الأحداث: | Date Created: 20220910 Date Completed: 20220913 Latest Revision: 20230701 |
| رمز التحديث: | 20230701 |
| مُعرف محوري في PubMed: | PMC10283041 |
| DOI: | 10.1007/978-1-0716-2712-9_7 |
| PMID: | 36087201 |
| قاعدة البيانات: | MEDLINE |
| تدمد: | 1940-6029 |
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| DOI: | 10.1007/978-1-0716-2712-9_7 |