|
| Status |
Public on Dec 31, 2017 |
| Title |
157 KO |
| Sample type |
SRA |
| |
|
| Source name |
splenic B cells
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
genotype: Csnk2b-/- (KO)
tissue: spleen
age: 8 weeks
|
| Extracted molecule |
total RNA |
| Extraction protocol |
spleen was taken from sacrificed mice and smashed to obtain a cell suspensions. B cells were isolated using magnetic separation and total RNA was prepared using QIAGEN RNeasy kit (#74106)
Total RNA samples were processed according to Dynabeads® mRNA DIRECTTM Purification Kit (Ambion) to isolate mRNA. The template library was prepared using the Ion Total RNA-Seq Kit v2 (Life Technologies). Quantity and size distribution of the library were analyzed using the Agilent Bioanalyzer 2100 DNA HS chip. Emulsion PCR using 10microl of 100pM library was performed using an OneTouch 2 instrument (Life Technologies) with an Ion PI Template OT2 200 kit following the manufacturer’s instructions. The enrichment of template library was achieved using the Ion OneTouch ES enrichment system (Life Technologies). Ion Proton sequencer and IPv2 chip were prepared according to the manufacturer’s recommendations.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Ion Torrent Proton |
| |
|
| Description |
AGM
|
| Data processing |
Reads preposessing was performed using cutadapt to remove adapter sequence. The minimum sequence length was set to 16
Reads were aligned on the reference genome using a two step procedures: at first reads were aligned using STAR program version 2.3.0e . The unmapped reads were aligned using bowtie2 version 2.2.2.1 setting the flag --local and --very-sensitive-local
Gene quantification was performed using htseq-count
raw counts were adjust for GC-content effect and to account for the different depth of sequencing usingfull quantile normalization implemented in EdaSeq R package
Differentially expressed genes were found using EdgeR R package setting the p-value < 0.05 after FDR correction
Genome_build: mm10
Supplementary_files_format_and_content: tab delimited text file containing the normalized reads counts for all the samples.
|
| |
|
| Submission date |
Oct 24, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Nicola Vitulo |
| E-mail(s) |
nicola.vitulo@univr.it
|
| Organization name |
University of Verona
|
| Department |
Department Biotechnology
|
| Street address |
Ca' Vignal 1, Strada Le Grazie 15
|
| City |
Verona |
| State/province |
Verona |
| ZIP/Postal code |
37134 |
| Country |
Italy |
| |
|
| Platform ID |
GPL18635 |
| Series (1) |
| GSE89082 |
Analysis of gene expression changes in B cells upon Csnk2b conditional deletion |
|
| Relations |
| BioSample |
SAMN05935560 |
| SRA |
SRX2265348 |
