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dc.contributor.authorGao, Fan
dc.contributor.authorWang, Kai
dc.date.accessioned2015-06-29T16:49:55Z
dc.date.available2015-06-29T16:49:55Z
dc.date.issued2015-05
dc.date.submitted2014-05
dc.identifier.issn1472-6750
dc.identifier.urihttp://hdl.handle.net/1721.1/97559
dc.description.abstractBackground As one of the genetic mechanisms for adaptive immunity, V(D)J recombination generates an enormous repertoire of T-cell receptors (TCRs). With the development of high-throughput sequencing techniques, systematic exploration of V(D)J recombination becomes possible. Multiplex PCR has been previously developed to assay immune repertoire; however, the use of primer pools leads to inherent biases in target amplification. In our study, we developed a “single-primer" ligation-anchored PCR method that may amplify the repertoire with much less biases. Results By utilizing a universal primer paired with a single primer targeting the conserved constant region, we amplified TCR-beta (TRB) variable regions from total RNA extracted from blood. Next-generation sequencing libraries were then prepared for Illumina HiSeq 2500 sequencer, which generates 151-bp read length to cover the entire V(D)J recombination region. We evaluated this approach on blood samples from healthy donors and from patients with malignant and benign meningiomas. Mapping of sequencing data showed that 64% to 96% of mapped TCRV-containing reads belong to TRB subtype. An increased usage of specific V segments and V-J pairing were observed in malignant meningiomas samples. The CDR3 sequences of the expanded V-J pairs were distinct in each malignant individual, even for pairing of TRBV7-3 with TRBJ2-2 that showed increased usage in both cases. Conclusions We demonstrated the technical feasibility and effectiveness of ligation-anchored PCR approach in capturing the TCR-beta landscapes. Further development of this technology may enable a comprehensive delineation of immune repertoire, including other forms of TCRs as well as immunoglobulins.en_US
dc.description.sponsorshipNational Human Genome Research Institute (U.S.) (Grant R01 HG006465)en_US
dc.description.sponsorshipAmerican Cancer Society (IRG-58-007-51)en_US
dc.publisherBioMed Centralen_US
dc.relation.isversionofhttp://dx.doi.org/10.1186/s12896-015-0153-9en_US
dc.titleLigation-anchored PCR unveils immune repertoire of TCR-beta from whole blooden_US
dc.typeArticleen_US
dc.identifier.citationGao, Fan, and Kai Wang. “Ligation-Anchored PCR Unveils Immune Repertoire of TCR-Beta from Whole Blood.” BMC Biotechnol 15, no. 1 (May 28, 2015).en_US
dc.contributor.departmentPicower Institute for Learning and Memoryen_US
dc.contributor.mitauthorGao, Fanen_US
dc.relation.journalBMC Biotechnologyen_US
dc.eprint.versionFinal published versionen_US
dc.type.urihttp://purl.org/eprint/type/JournalArticleen_US
eprint.statushttp://purl.org/eprint/status/PeerRevieweden_US
dc.date.updated2015-06-29T08:39:17Z
dc.language.rfc3066en
dc.rights.holderGao and Wang; licensee BioMed Central.
dspace.orderedauthorsGao, Fan; Wang, Kaien_US
mit.licenseOPEN_ACCESS_POLICYen_US
mit.metadata.statusComplete


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