Hanlee P. Ji (Stanford University School of Medicine)

한빛사논문

조회54

Hanlee P. JiHanlee P. Ji

Stanford University School of Medicine

Proc. Natl. Acad. Sci. USA, Jul 30 2024, 121 (31) | https://doi.org/10.1073/pnas.2322834121 Resolving the 22q11.2 deletion using CTLR-Seq reveals chromosomal rearrangement mechanisms and individual variance in breakpoints

Bo Zhou^a,b,2,1, Carolin Purmann^a,b,c,1, Hanmin Guo^a,b,c,d,e,1, GiWon Shin^f,3, Yiling Huang^a,c, Reenal Pattni^a,c, Qingxi Meng^f, Stephanie U. Greer^f, Tanmoy Roychowdhury^g,4, Raegan N. Wood^f, Marcus Ho^a,c, Heinrich zu Dohna^h, Alexej Abyzov^g, Joachim F. Hallmayer^a, Wing H. Wong^d,e,2, Hanlee P. Ji^f,2, and Alexander E. Urban^a,b,c,i,2

^aDepartment of Psychiatry and Behavioral Sciences, Stanford University School of Medicine, Stanford, CA 94305

^bStanford Maternal and Child Health Research Institute, Stanford University School of Medicine, Stanford, CA 94305

^cDepartment of Genetics, Stanford University School of Medicine, Stanford, CA 94305

^dDepartment of Statistics, Stanford University, Stanford, CA 94305

^eDepartment of Biomedical Data Science, Stanford University School of Medicine, Stanford, CA 94305

^fDivision of Oncology, Department of Medicine, Stanford University School of Medicine, Stanford, CA 94305

^gDivision of Computational Biology, Department of Quantitative Health Sciences, Mayo Clinic, Rochester, MN 55905

^hDepartment of Biology, American University of Beirut, Beirut 1107 2020, Lebanon

ⁱProgram on Genetics of Brain Function, Stanford Center for Genomics and Personalized Medicine, Stanford University School of Medicine, Stanford, CA 94305

¹B.Z., C.P., and H.G. contributed equally to this work.

²To whom correspondence may be addressed. : Wing H. Wong, Hanlee P. Ji, Alexander E. Urban

³Present address: McDonnell Genome Institute, Washington University School of Medicine, Saint Louis, MO 63108.

⁴Present address: Department of Genetics, Yale School of Medicine, New Haven, CT 06510.

Abstract

We developed a generally applicable method, CRISPR/Cas9-targeted long-read sequencing (CTLR-Seq), to resolve, haplotype-specifically, the large and complex regions in the human genome that had been previously impenetrable to sequencing analysis, such as large segmental duplications (SegDups) and their associated genome rearrangements. CTLR-Seq combines in vitro Cas9-mediated cutting of the genome and pulse-field gel electrophoresis to isolate intact large (i.e., up to 2,000 kb) genomic regions that encompass previously unresolvable genomic sequences. These targets are then sequenced (amplification-free) at high on-target coverage using long-read sequencing, allowing for their complete sequence assembly. We applied CTLR-Seq to the SegDup-mediated rearrangements that constitute the boundaries of, and give rise to, the 22q11.2 Deletion Syndrome (22q11DS), the most common human microdeletion disorder. We then performed de novo assembly to resolve, at base-pair resolution, the full sequence rearrangements and exact chromosomal breakpoints of 22q11.2DS (including all common subtypes). Across multiple patients, we found a high degree of variability for both the rearranged SegDup sequences and the exact chromosomal breakpoint locations, which coincide with various transposons within the 22q11.2 SegDups, suggesting that 22q11DS can be driven by transposon-mediated genome recombination. Guided by CTLR-Seq results from two 22q11DS patients, we performed three-dimensional chromosomal folding analysis for the 22q11.2 SegDups from patient-derived neurons and astrocytes and found chromosome interactions anchored within the SegDups to be both cell type-specific and patient-specific. Lastly, we demonstrated that CTLR-Seq enables cell-type specific analysis of DNA methylation patterns within the deletion haplotype of 22q11DS.

논문정보

형식Research article
게재일2024년 07월 (BRIC 등록일 2024-07-31)
연구진국외 연구진
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