|
| Status |
Public on Aug 07, 2023 |
| Title |
cKO Input 71320 |
| Sample type |
SRA |
| |
|
| Source name |
Neocortex
|
| Organism |
Mus musculus |
| Characteristics |
tissue: Neocortex
genotype: cKO
fraction: Input
|
| Growth protocol |
Wild type (WT) littermate control and Celf4 conditional-deletion animals were generated by crossing Celf4f/f (B6.129-Celf4tm1.1Frk/Frk (Celf4f/f mice; Jax strain: 018126) from Jackson Laboratory) mice with B6.129S2-Emx1tm1(cre)Krj/J (Emx1-Cre mice, Jax strain: 005628) transgenic line that drives the expression of Cre recombinase in the cortical progenitors and postmitotic neurons of neocortex.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Mouse embryonic neocortices at postnatal day 0 (P0) were dissected in ice-cold HBSS media supplemented with 5mg/ml glucose and 20 mM HEPES (pH 7.2), followed by freezing on dry ice. Three mouse cortices were pooled and homogenized in freshly prepared polysome extraction buffer supplemented with EDTA-free protease inhibitor, RNaseOut, DTT, and cycloheximide. Lysis was performed by pipetting gently up and down on ice for 15 minutes. Lysate was centrifuged at 5k rpm for 10 min at 4C and the resulting supernatant was spun at 14k for 5 min at 4C. Following quantification, 120 ug neocortical input was layered onto 10-50% sucrose density gradients in thin wall polypropylene tubes. A portion (25%) was stored for processing as input. Gradients were spun at 39K rpm at 4C for 120 min. Polysome profiles were generated with a continuous flow Fluorinert FC-40 and A254 recording with a Brandel fraction collector with UV absorbance recorder. Samples were frozen at -80C until use. Pooled monosomes (40S-60S-80S) and polysomes fractions were pooled for isolation of RNA by extraction with TRIzol LS. RNA was precipitated with one volume of isopropanol/glycogen, then washed with 75% ethanol and air-dried. Following resuspension, genomic DNA was destroyed with DNase treatment.
Illumina TruSeq RNA Library Kit
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
071320-Emx-Cre-Celf4-cKO-Input
|
| Data processing |
Reads trimmed and filtered with fastp v0.12.2
Assembled with Ensembl GRCm39 cDNA index using Kallisto v0.46.0
Counts extracted with tximport
Counts imported to DESeq2
Assembly: GRCm39
Supplementary files format and content: Tab-delimited normalized counts per sample per Ensembl transcript ID exported from DESeq2
|
| |
|
| Submission date |
Sep 28, 2022 |
| Last update date |
Aug 07, 2023 |
| Contact name |
Ronald P. Hart |
| E-mail(s) |
rhart@rutgers.edu
|
| Phone |
848-445-1783
|
| Organization name |
Rutgers University
|
| Department |
Cell Biology & Neuroscience
|
| Street address |
604 Allison Rd Rm B430
|
| City |
Piscataway |
| State/province |
NJ |
| ZIP/Postal code |
08854 |
| Country |
USA |
| |
|
| Platform ID |
GPL17021 |
| Series (2) |
| GSE214328 |
Mouse Celf4 cKO fetal neocortex polysome fractions |
| GSE214534 |
The role of mRNA translation in the synaptic development of prenatal mammalian neocortex |
|
| Relations |
| BioSample |
SAMN31062413 |
| SRA |
SRX17732095 |
