|
| Status |
Public on Dec 14, 2020 |
| Title |
erk7 starved rep2 |
| Sample type |
SRA |
| |
|
| Source name |
whole larvae
|
| Organism |
Drosophila melanogaster |
| Characteristics |
genotype: erk7 mutant
tissue: whole larvae
developmental stage: 3rd instar larvae
|
| Growth protocol |
Control and ERK7 mutant flies were kept in egg laying champers with apple juice plates and yeast paste for 3 hours. 24 hours later the 1st instar larvae were transferred to standard lab food. 48 hours later larvae were transferred to either PBS + 0.5% agarose food or a fresh standard lab food. 6 hours later, larvae were collected from both food types for RNA extraction.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Larvae were frozen in liquid nitrogen and homogenized with a pestle and mortar in lysis buffer and vortexed. Samples were frozen for complete lysis of the tissue. Total RNA was extracted with Nucleospin RNA II kit according to manufacturer's protocol. Elution was done with water and the samples stored at -80°C. Library construction was performed by DNA sequencing and genomics lab at the Institute of Biotechnology with the following protocol: the total RNA samples were first treated with Dnase I. Then the mRNA was enriched by using the oligo(dT) magnetic beads. Mixed with the fragmentation buffer, the mRNA was fragmented into short fragments. The first stand of cDNA was synthesized by using random hexamer-primer. Buffer, dNTPs, RNase H and DNA polymerase I were added to synthesize the second strand. The double strand cDNA was purified with magnetic beads. End reparation and 3'-end single nucleotide A addition was performed. Finally, sequencing adaptors were ligated to the fragments and fragment enrichment was done by PCR amplification.
Library construction was performed by DNA sequencing and genomics lab at the Institute of Biotechnology with the following protocol: mRNA was enriched by using the oligo(dT) magnetic beads. Mixed with the fragmentation buffer, the mRNA was fragmented into short fragments. The first stand of cDNA was synthesized by using random hexamer-primer. Buffer, dNTPs, RNase H and DNA polymerase I were added to synthesize the second strand. The double strand cDNA was purified with magnetic beads. End reparation and 3'-end single nucleotide A addition was performed. Finally, sequencing adaptors were ligated to the fragments and fragment enrichment was done by PCR amplification.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
all_comparisons.txt
S 14
|
| Data processing |
Sequenced reads were trimmed with Trimmomatic with parameters ILLUMINACLIP:TruSeq3-SE:2:30:10 LEADING:20 TRAILING:20 SLIDINGWINDOW:4:15 MINLEN:36
Sequences were mapped using TopHat (v2.1.0) to dmel R6.10 with parameter -p 4 and quantified with HTSeq (v2.7.6) with parameters -m HTSeq.scripts.count -m union -s reverse -a 10 -t exon -i Parent
Differentially expressed genes were calculated with R package limma with Benjamini-Hochberg correction, with genes with low counts (cpm<1 in more than one replicate) filtered
Genome_build: Flybase Drosophila melanogaster R6.10
Supplementary_files_format_and_content: tab delimited.txt file includes read counts for expressed genes for all samples and their statistics
|
| |
|
| Submission date |
Dec 16, 2018 |
| Last update date |
Dec 14, 2020 |
| Contact name |
Krista Kokki |
| Organization name |
University of Helsinki
|
| Department |
Department of Biosciences (Genetics)
|
| Lab |
Hietakangas lab
|
| Street address |
Viikinkaari 9, Biocenter 1, Institute of Biotechnology P.O.Box. 56
|
| City |
Helsinki |
| ZIP/Postal code |
00014 |
| Country |
Finland |
| |
|
| Platform ID |
GPL19132 |
| Series (1) |
| GSE123901 |
Coordinated control of adiposity and growth by anti‐anabolic kinase ERK7 |
|
| Relations |
| BioSample |
SAMN10600716 |
| SRA |
SRX5137749 |
