USC Stem Cell Seminar: Mo Li, King Abdullah Univ. of Science & Technology—"Nanopore sequencing resolving CRISPR-induced mutations in human pluripotent stem cells and germline mitochondria genetic heterogeneity at the single-molecule and single-cell level"

Tuesday, July 27 at 11:00am to 12:00pm

This is a past event.
Virtual Event

For Zoom links for the seminar and discussions, please email Belen Lopez at belen.lopez@med.usc.edu.

Genome editing by CRISPR-Cas9 can lead to large deletions and complex rearrangements in mouse embryonic stem cells (mESCs). It is unclear if this phenomenon also happens in human ESCs (hESCs), and, more importantly, an unbiased and quantitative characterization of CRISPR-induced mutagenesis is lacking due to limitations of current strategies. We develop individual molecular sequencing (IDMseq) for base-resolution haplotype-resolved quantitative characterization of diverse types of rare variants using long-read sequencing platforms. It provides the first quantitative evidence of persistent nonrandom large structural variants and an increase in SNVs at the on-target locus following repair of double-strand breaks induced by CRISPR-Cas9 in hESCs. We further applied the principle of IDMseq to study the mtDNA genetic heterogeneity. The ontogeny and dynamics of mtDNA heteroplasmy remain unclear due to limitations of current mtDNA sequencing methods. We developed individual Mitochondrial Genome sequencing (iMiGseq) of full-length mtDNA for ultra-sensitive variant detection, complete haplotyping, and unbiased evaluation of heteroplasmy levels. iMiGseq uncovers unappreciated levels of heteroplasmic variants in single healthy human oocytes well below the conventional detection limit. Many detected rare variants are deleterious and associated with diseases. iMiGseq revealed dramatic shifts in variant frequency and clonal expansion of large structural variants of mtDNA during oogenesis in healthy humans and mice. iMiGseq provided haplotype-resolved SNV and large structural variants of mtDNA in NARP/Leigh syndrome patient-derived iPSCs. Therefore, iMiGseq could not only elucidate the mitochondrial etiology of diseases, but also help diagnose and prevent mitochondrial diseases with unprecedented precision.

Host: Zhongwei Li

Event Type

Lecture / Talk / Workshop

Campus

Other Location

Tags

stemcell

Department
Stem Cell at USC
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