|
| Status |
Public on Jan 25, 2019 |
| Title |
KO Bone-1 |
| Sample type |
SRA |
| |
|
| Source name |
Lin-Sca1+cKIT+CD48-CD150+
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57/Bl6
tissue: Bone marrow - Bone fraction
genotype: Gfra2 KO
|
| Treatment protocol |
no treatment
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Bone & marrow fraction were separated from tibia, femur and humeri of WT and Gfra2KO mice. After RBC lysis, mature hematopoietic cells were depleted using the EasyStep Mouse Hematopoietic Progenitor Cell Isolation Cocktail (Stem cell Technologies, 19756) according to the manufacturer’s protocol. The depleted cell population was stained for 20 minutes in PBS with 2 % FCS for LSK CD48-CD150+ markers. After dead cell exclusion using DAPI, pools of 30 viable LSK CD48-CD150+ cells were sorted using BD Influx™ cell sorter into 4 µl lysis buffer (0.5 U/µl SUPERase In RNase Inhibitor in 0.2 % (v/v) Triton-X-100) containing 12.5 mM DTT and 2.5 mM dNTP. RNAseq was performed following Smart-seq2 protocol (Picelli 2014)
Pooled libraries of 4 replicates per sample were prepared using the Illumina Nextera XT DNA preparation kit & sequencing was performed on the Illumina HiSeq 4000
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Data processing |
RNA-seq reads were aligned to mm10 using GSNAP (version 2015-09-29) with parameters (-B 5 -t 24 -n 1 -Q -N 1).
Reads in features were counted with htseq-count (HTSeq version 0.5.3p3) with the parameter (-s no).
Genome_build: mm10
Supplementary_files_format_and_content: HTSeq counts (.txt)
|
| |
|
| Submission date |
Jan 25, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Evangelia Diamanti |
| E-mail(s) |
ed347@cam.ac.uk
|
| Phone |
01223 62317
|
| Organization name |
University of Cambridge
|
| Department |
Haematology
|
| Lab |
Gottgens
|
| Street address |
Wellcome Trust / MRC Building, Hills Road
|
| City |
Cambridge |
| ZIP/Postal code |
CB2 0XY |
| Country |
United Kingdom |
| |
|
| Platform ID |
GPL21103 |
| Series (1) |
| GSE94078 |
Gene expression profiling of LSKSLAM cells in mice deficient for cholinergic fibers |
|
| Relations |
| BioSample |
SAMN06271530 |
| SRA |
SRX2517795 |
