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Sample GSM3680565 Query DataSets for GSM3680565
Status Public on Mar 20, 2019
Title AAV1
Sample type SRA
 
Source name Retina
Organism Mus musculus
Characteristics sample type: P21 retinae
treatment: intravitreally injected with AAV-Cas9-VQR and AAV-U6sgRNAmMUT-RHOp-GFP-t2a-rtTA PHP.B serotype
background strain: Heterozygous P23H knock-in mice (The Jackson Laboratory, Bar Harbor, ME)
Treatment protocol Electoporated retinae were dissected under a fluorescence dissecting microscope. GFP+ retinae were selected for further analysis based on fluorescence intensity and diffusion but also retinal integrity.
Extracted molecule genomic DNA
Extraction protocol In case of indel analysis, retinae were dissociated (papain 10 U/ml, Worthington, 10 min, 37°C) (68) and FACS-sorted (MoFlo, Beckman Coulter, Inc.).
Genomic PCR products were subjected to library preparation using the NEBNext Ultra II DNA library Kit (NEB). Briefly, common primers are added by ligation, following by limited number of PCR cycles (eight cycles) where specific barcode for each sample was added. Amplicones were purificated with AMPure beads (Beckman Coulter) and run onto Bioanalyzer 2100 (Agilent). Equimolar amounts of library were mixed, diluited and sequenced with an Illumina MiSeq system. CASAVA software (Illumina) was used for demultiplexing.
 
Library strategy OTHER
Library source genomic
Library selection other
Instrument model Illumina MiSeq
 
Data processing Fastq reads were aligned to the mm10 reference genome with Bowtie.
NHEJ as well as frameshift/in-frame mutations were quantified with CRISPResso.
Genome_build: mm10
 
Submission date Mar 20, 2019
Last update date Mar 21, 2019
Contact name Luca Massimino
E-mail(s) admin@lucamassimino.com
Phone 3389039500
Organization name Ospedale San Raffaele
Department Gastroenterology
Street address Via Olgettina, 58
City Milano
State/province MI
ZIP/Postal code 20100
Country Italy
 
Platform ID GPL16417
Series (1)
GSE128579 Cas9/sgRNA selective targeting of the P23H Rhodopsin mutant allele for treating retinitis pigmentosa by intravitreal AAV9.PHP.B-based delivery
Relations
BioSample SAMN11176072
SRA SRX5546878

Supplementary file Size Download File type/resource
GSM3680565_AAV1_S2_mm10.bedGraph.gz 632 b (ftp)(http) BEDGRAPH
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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