SMRT Sequencing for Parallel Analysis of Multiple Targets and Accurate SNP Phasing.
Single-molecule real-time (SMRT) sequencing generates much longer reads than other widely used next-generation (next-gen) sequencing methods, but its application to whole genome/exome analysis has been limited. Here, we describe the use of SMRT sequencing coupled with barcoding to simultaneously analyze one or a small number of genomic targets derived from multiple sources. In the budding yeast system, SMRT sequencing was used to analyze strand-exchange intermediates generated during mitotic recombination and to analyze genetic changes in a forward mutation assay. The general barcoding-SMRT approach was then extended to diffuse large B-cell lymphoma primary tumors and cell lines, where detected changes agreed with prior Illumina exome sequencing. A distinct advantage afforded by SMRT sequencing over other next-gen methods is that it immediately provides the linkage relationships between SNPs in the target segment sequenced. The strength of our approach for mutation/recombination studies (as well as linkage identification) derives from its inherent computational simplicity coupled with a lack of reliance on sophisticated statistical analyses.
Duke Scholars
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- Yeasts
- Reproducibility of Results
- Recombination, Genetic
- Polymorphism, Single Nucleotide
- Mutation
- Lymphoma
- Humans
- High-Throughput Nucleotide Sequencing
- Genomics
- Genetic Linkage
Citation
Published In
DOI
EISSN
Publication Date
Volume
Issue
Start / End Page
Location
Related Subject Headings
- Yeasts
- Reproducibility of Results
- Recombination, Genetic
- Polymorphism, Single Nucleotide
- Mutation
- Lymphoma
- Humans
- High-Throughput Nucleotide Sequencing
- Genomics
- Genetic Linkage