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Sample GSM2402793 Query DataSets for GSM2402793
Status Public on Dec 16, 2016
Title SP-Itgam_Itgam.sca
Sample type SRA
 
Source name ~4,000 bulk-sorted mouse ex-vivo myeloid cells
Organism Mus musculus
Characteristics strain: transgenic Cas9-GFP C57BL/7
age: 6 to 8 weeks-old
tissue: hind leg bones
tissue: bone marrow
selection marker: GFP+BFP+CD11c+
treatment: unstimulated
grna in the infecting vector: Itgam-BFP x Cebpb-mCherry
Treatment protocol Cells were infected on day 2 post plating with lentivirus vectors, each containing one gRNA and one fluorescent selection marker. On day 7, cells were challenged with LPS for 4 hours before harvest.
Growth protocol Bone marrows were flushed from hind leg bones from euthanized 6 to 8 weeks-old mice housed at the Weizmann Institute animal facility. Cell suspensions were obtained, red blood cells lysed, and cells grown on 6-well petri dishes on media supplemented with GM-CSF.
Extracted molecule genomic DNA
Extraction protocol Cells were scrapped from the plate and stained with FACS markers antibodies as indicated above (selection marker).
3' end mRNA libraries were prepared for sequencing using CRISP-seq (Jaitin et al., Cell 2016), which includes a module of MARS-seq (Jaitin et al., Science 2014) and a module of UGI-seq.
single cell RNA-seq and UGI-seq for gene expression quantitation and gRNA identification, respectively (paired-end + i7 for sample index (7 bases))
 
Library strategy OTHER
Library source genomic
Library selection other
Instrument model Illumina MiSeq
 
Description Well with cells infected with a mix of Itgam-gRNA BFP and Cebpb-gRNA mCherry lentiviruses
Sample 54
Data processing bcl2fastq/2.15.0.4
Sequences with RMT of low quality (defined as RMT with minimum Phred score of less than 27) were filtered out.
Pool-barcode and well-barcode-RMT were extracted from the first and second end of the read (respectively) and concatenated to the fastq header, delimited by a underscore i.e. POOL_BARCODE_WELL_BARCODE_RMT while "NNNNNN" was used as a place holders if plate barcode was not used.
Reads were separated by POOL_BARCODE_WELL_BARCODE header data, allowing 1 sequencing error. This process created a single fastq file for each source well.
Genome_build: mm9
Supplementary_files_format_and_content: lists of gene counts; specific amplicons within genomic DNA;
Supplementary_files_format_and_content: CIGAR string as defined in SAM file sprcifications, showing count of uniq strings. This is used to estimate the editing percentage of the target loci.
 
Submission date Nov 24, 2016
Last update date May 15, 2019
Contact name Ido Amit
E-mail(s) ido.amit@weizmann.ac.il
Phone 972-8-9343338
Organization name Weizmann Institute of Science
Department Immunology
Street address 234 Herzl st.
City Rehovot
ZIP/Postal code 760001
Country Israel
 
Platform ID GPL16417
Series (2)
GSE90487 Dissecting immune circuits by linking CRISPR pooled screens with single cell RNA-seq [SET2]
GSE90488 Dissecting immune circuits by linking CRISPR pooled screens with single cell RNA-seq
Relations
BioSample SAMN06053562
SRA SRX2373901

Supplementary file Size Download File type/resource
GSM2402793_SP-Itgam_Itgam.sca.txt.gz 17.4 Kb (ftp)(http) TXT
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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