|
| Status |
Public on Dec 15, 2021 |
| Title |
Enterobacteriaceae_Gnotobiotic pupae rep3 [ent.pup6] |
| Sample type |
SRA |
| |
|
| Source name |
Enterobacteriaceae_pupae
|
| Organism |
Aedes aegypti |
| Characteristics |
treatment: Enterobacteriaceae
Stage: Pupae
|
| Treatment protocol |
Axenic larvae were made gnotobiotic larvae by adding to the flask 5.10E5 CFU/ml of selected bacterial isolates.
|
| Growth protocol |
Aedes aegypti eggs were surface sterilized and were allowed to hatch in sterile water in a vacuum chamber, then were transferred to sterile 25-ml tissue-culture flasks with filter-top lids and maintained in 15 ml of sterile water. Larvae were maintained on 60 μl of sterile fish food (autoclaved food for 20 min) every other day.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Pool of 12 axenic larvae, L4 larvae, pupae and adult mosquitoes were surface sterilized and total RNA was extracted using Trizol.
Axenic larva, L4 larva, pupa and adult mosquito libraries for controls (non-axenics) and for gnotobiotics (Flavobacterium, Enterobacteriaceae, Lysobacter and Paenibacillus treatments) were prepared from total RNA of pool of 12 individuals. Removal of ribosomal RNA, fragmentation and library preparation were performed by using the TruSeq Stranded mRNA library preparation kit (Illumina) following manufacturer’s instructions. Libraries quality was checked on a DNA1000 Bioanalyzer chip (Agilent) and quantification made by QuBit DNA HS kit (ThermoFisher). single reads of 65 nucleotides in length (à verifier) were generated on a HiSeq2500 sequencing platform (Illumina).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
entero6_S10_L002
|
| Data processing |
Reads were cleaned of adapter sequences and low-quality sequences using cutadapt version 1.11.Only sequences ≥25 nucleotides (nt) in length were considered for further analysis.
STAR version 2.5.0a, with default parameters, was used for alignment on the Ae. aegypti genome
Genes were counted using featureCounts version 1.4.6-p35from Subreads package (parameters: -t exon, -g gene_id and -s 1)
Count data were analyzed using R version 3.6.1 and the Bioconductor package DESeq2 version 1.24.0. The normalization and dispersion estimation were performed with DESeq2 using the default parameter and statistical tests for differential expression were performed applying the independent filtering algorithm. A generalized linear model was set in order to test for the differential expression between gnotobiotic and non-axenic for larval, pupal and adult stages. For each pairwise comparison, raw p-values were adjusted for multiple testing according to the Benjamini and Hochberg (BH) procedure and genes with both an absolute log2FC > 1 and an adjusted p-value lower than 0.05 were considered differentially expressed.
Genome_build: AaegL5.3, http://vectorbase.org
Supplementary_files_format_and_content: Count matrix from DESeq2
|
| |
|
| Submission date |
Apr 28, 2021 |
| Last update date |
Dec 15, 2021 |
| Contact name |
Rachel Legendre |
| E-mail(s) |
rachel.legendre@pasteur.fr
|
| Organization name |
Institut Pasteur
|
| Department |
Research and Resource Center for Scientific Informatics
|
| Lab |
Hub of Bioinformatics and Biostatistics
|
| Street address |
28, rue du docteur Roux
|
| City |
Paris |
| ZIP/Postal code |
75724 |
| Country |
France |
| |
|
| Platform ID |
GPL19664 |
| Series (1) |
| GSE173472 |
Transcriptomic analysis of gnotobiotic Aedes aegypti mosquitoes |
|
| Relations |
| BioSample |
SAMN18901859 |
| SRA |
SRX10695010 |
