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Series GSE146998 Query DataSets for GSE146998
Status Public on Sep 01, 2020
Title Indel rate analysis:Start codon disruption with CRISPR/Cas9 prevents murine Fuchs' endothelial corneal dystrophy
Organism Mus musculus
Experiment type Other
Summary A missense mutation of collagen type VIII alpha 2 chain (COL8A2) gene leads to early onset Fuchs’ endothelial corneal dystrophy (FECD), which can cause blindness through progressive loss of corneal endothelial cells. We established a novel procedure for achieving structural and functional rescue of the post-mitotic corneal endothelium without surgery, using CRISPR/Cas9-based postnatal gene editing in a mouse model of FECD. A single intraocular injection of an adenovirus encoding both the Cas9 gene and guide RNA (Ad-Cas9-Col8a2gRNA) at a titer below the cytotoxic threshold, efficiently knocked down COL8A2 protein expression in corneal endothelial cells, prevented endothelial cell loss, and rescued corneal endothelium pumping function in adult Col8a2 mutant mice. Here, to determine the indel rate in mouse corneal endothelium, we performed deep sequencing of PCR products (including the target site) amplified from genomic DNA of corneal endothelium. We found that the indel rate was 23.7 ± 4.5% in mouse corneal endothelium. Most insertions were 1bp insertions (19.8 ± 4.0% in total reads), while 2bp deletions were the most frequent (1.0 ± 0.3% in total reads). We moreover found that A or T insertion was predominant, with the proportion of A:T:G:C being 48.7 : 44.6 : 1.8 : 4.9.
Overall design gRNA target site was amplified by PCR, and indel rate was determined by deep sequencing
Contributor(s) Uehara H, Ambati B
Citation(s) 34100716
Submission date Mar 15, 2020
Last update date Jul 02, 2021
Contact name Hironori Uehara
Organization name Loma Linda University
Department Ophthalmology
Lab Ambati lab
Street address 11021 Campus st
City Loma Linda
State/province California
ZIP/Postal code 92350
Country USA
Platforms (1)
GPL16417 Illumina MiSeq (Mus musculus)
Samples (4)
GSM4411938 15580X1 Cornea rep1
GSM4411939 15580X2 Cornea rep2
GSM4411940 15580X3 Cornea rep3
This SubSeries is part of SuperSeries:
GSE146999 Start codon disruption with CRISPR/Cas9 prevents murine Fuchs' endothelial corneal dystrophy
BioProject PRJNA612702
SRA SRP252871

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Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE146998_15580R_IndelRate_MsCornealEndothelium.xlsx 35.8 Kb (ftp)(http) XLSX
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Raw data are available in SRA
Processed data are available on Series record

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