|
| Status |
Public on Oct 02, 2018 |
| Title |
0h_1 |
| Sample type |
SRA |
| |
|
| Source name |
pupae (0h after puparium formation)
|
| Organism |
Drosophila melanogaster |
| Characteristics |
organ: Brain
selection marker: DsRed +
tissue: Isolated γ neurons
developmental stage: pupae (0h after puparium formation)
strain: y,w/+; UAS-RedStinger/+;71G10/+
|
| Growth protocol |
All fly strains were reared under standard laboratory conditions at 25°C on molasses containing food. Males and females were chosen at random. Developmental stage is referred to in the relevant places while adult refers to 3-5 days post eclosion.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Brains were dissected a cold Ringer’s solution, incubated with collagenase/dispase mix at 29ºC, washed in dissociation solution, mechanically dissociated into single cells and transferred via 35um mesh. 1000 DsRed+ positive cells were sorted directly into 100μl Pico-Pure RNA isolation kit extraction buffer followed by RNA extraction using the kit or stored in -80 for later use.
Library constracted using MARS-seq technique (Jaitin et al, Science 2014). Briefly mRNA was barcoded, converted into cDNA and linearly amplified by T7 in vitro transcription.
single end protocol ( samples used read2 to read molecule barcodes and UMI)
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Illumina bcl2fastq software used for basecalling.
Sample barcodes were extracted from read 2 and concatenated to the fastq header of read 1. Barcode is of size 7 followed by UMI of size 8
alignment: hisat 0.1.5 with deafult parameters
filter PCR amplification bias using alignment break site and UMI barcode (size 8).
create tag directory using makeTagDirectory from HOMER tools (http://homer.salk.edu/homer/ngs/)
HOMER, quantifying RNA expression in genes: analyzeRepeats.pl DM6.gtf none -d [TAG DIR] -count 3utr -condenseGenes -strand +
Genome_build: DM6
Supplementary_files_format_and_content: tab-delimited text files include mRNA molecule count values for each Sample
|
| |
|
| Submission date |
Jul 27, 2017 |
| 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 |
GPL19132 |
| Series (1) |
| GSE101946 |
RNA-seq - Combining developmental and perturbation-seq uncovers transcriptional modules orchestrating neuronal remodeling |
|
| Relations |
| BioSample |
SAMN07418819 |
| SRA |
SRX3041860 |
