A new hybrid approach for MHC genotyping: high-throughput NGS and long read MinION nanopore sequencing, with application to the non-model vertebrate Alpine chamois (Rupicapra rupicapra)

dc.creatorFuselli, S.
dc.creatorBaptista, R. P.
dc.creatorPanziera, A.
dc.creatorMagi, A.
dc.creatorGuglielmi, S.
dc.creatorTonin, R.
dc.creatorBenazzo, A.
dc.creatorBauzer, L. G.
dc.creatorMazzoni, C. J.
dc.creatorBertorelle, G.
dc.date2018-09-28T13:43:36Z
dc.date2018-09-28T13:43:36Z
dc.date2018
dc.date.accessioned2023-09-26T20:39:35Z
dc.date.available2023-09-26T20:39:35Z
dc.identifierFUSELLI, S. et al. A new hybrid approach for MHC genotyping: high-throughput NGS and long read MinION nanopore sequencing, with application to the non-model vertebrate Alpine chamois (Rupicapra rupicapra). Heredity. v.121, p.293–303, 2018.
dc.identifier0018-067X
dc.identifierhttps://www.arca.fiocruz.br/handle/icict/29146
dc.identifier10.1038/s41437-018-0070-5
dc.identifier1365-2540
dc.identifier.urihttps://repositorioslatinoamericanos.uchile.cl/handle/2250/8861494
dc.descriptionThe major histocompatibility complex (MHC) acts as an interface between the immune system and infectious diseases. Accurate characterization and genotyping of the extremely variable MHC loci are challenging especially without a reference sequence. We designed a combination of long-range PCR, Illumina short-reads, and Oxford Nanopore MinION long-reads approaches to capture the genetic variation of the MHC II DRB locus in an Italian population of the Alpine chamois (Rupicapra rupicapra). We utilized long-range PCR to generate a 9 Kb fragment of the DRB locus. Amplicons from six different individuals were fragmented, tagged, and simultaneously sequenced with Illumina MiSeq. One of these amplicons was sequenced with the MinION device, which produced long reads covering the entire amplified fragment. A pipeline that combines short and long reads resolved several short tandem repeats and homopolymers and produced a de novo reference, which was then used to map and genotype the short reads from all individuals. The assembled DRB locus showed a high level of polymorphism and the presence of a recombination breakpoint. Our results suggest that an amplicon-based NGS approach coupled with single-molecule MinION nanopore sequencing can efficiently achieve both the assembly and the genotyping of complex genomic regions in multiple individuals in the absence of a reference sequence.
dc.description2030-01-01
dc.formatapplication/pdf
dc.languageeng
dc.publisherSpringer Nature
dc.rightsrestricted access
dc.subjectgenotipagem de MHC
dc.subjectabordagem híbrida
dc.subjectnanoporos MinION
dc.subjectRupicapra rupicapra
dc.subjectRupicapra rupicapra
dc.subjectMHC genotyping
dc.subjecthybrid approach
dc.subjectMinION nanopore sequencing
dc.titleA new hybrid approach for MHC genotyping: high-throughput NGS and long read MinION nanopore sequencing, with application to the non-model vertebrate Alpine chamois (Rupicapra rupicapra)
dc.typeArticle


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