|
| Status |
Public on Aug 31, 2024 |
| Title |
pooled whole embryos 210 ng/L repC |
| Sample type |
SRA |
| |
|
| Source name |
whole body tissue
|
| Organism |
Xenopus laevis |
| Characteristics |
age: 6 days post fertilization
exposure compound: lead (II) nitrate
treatment: 210 ng/L lead (II) nitrate
|
| Treatment protocol |
Embryos at approximately Nieuwkoop and Faber (NF) stage 29 (~48 h post-fertilization) were used for treatment and exposed for 96h. Embryos were placed in 50 mL plastic petri dishes containing one of the folllowing solutions of lead nitrate 0, 210 or 630 μg/L. During the exposure, animals were kept in an environmental chamber with ambient temperature of 21°C and a 16 h light: 8 h dark photoperiod.
|
| Growth protocol |
Embryos were obtained by breeding adult Xenopus laevis from an established colony. Adults were maintained at 18 ± 1°C water temperature and a 12 h light:12 h dark photoperiod. Males and female pairs were selected for breeding, and gradually acclimated to 21 ± 1°C. Mating was induced by injecting human chorionic gonadotropin (hCG, Sigma-Aldrich, St. Louis, MN) dissolved in sterilized phosphate buffered saline (PBS). Males and females were randomly paired for breeding, placed into aquaria with water temperature maintained at 21 ± 1 °C, and left to spawn overnight. After collection the following day, selection and preparation of embryos closely followed protocols outlined in “Standard guide for conducting the Frog Embryo Teratogenesis Assay-Xenopus (FETAX)” (ASTM, 2019) with some modifications. Up to 30 selected embryos were placed in each egg cup in an aquarium to incubate in facility water maintained at 21 ± 1 °C for 48 h. Animals used in this study were handled in accordance with the University of Saskatchewan’s Animal Research Ethics Board (protocol #20160090) and the Canadian Council on Animal Care guidelines for humane animal use.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Tissue was homogenized and RNA was extracted using the RNeasy Plus Universal Mini kit (QIAGEN) according to the manufacturer’s instructions.
Libraries were generated from 250 ng of total RNA as following: mRNA enrichment was performed using the NEBNext Poly(A) Magnetic Isolation Module (New England BioLabs). cDNA synthesis was achieved with the NEBNext RNA First Strand Synthesis and NEBNext Ultra Directional RNA Second Strand Synthesis Modules (New England BioLabs). The remaining steps of library preparation were done using and the NEBNext Ultra II DNA Library Prep Kit for Illumina (New England BioLabs). Adapters and PCR primers were purchased from New England BioLabs. Libraries were quantified using the Kapa Illumina GA with Revised Primers-SYBR Fast Universal kit (Kapa Biosystems). Average size fragment was determined using a LabChip GXII (PerkinElmer) instrument.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
SigGene_Medium_vs_Control_Hisat2
XL_lead_II_nitrate_HiSat2.tabular
XE3-MCP3-WT4
|
| Data processing |
The fastq files were trimmed using TrimGalore to remove adapter contamination and carry out quality control steps. This was carried out in galaxy.ecotoxxplorer.ca
The trimmed sequencing reads were aligned to the reference Xenopus laevis genome (GenBank assembly accession: GCA_001663975.1) using HiSat2 to obtain the raw count files. This was carried out in galaxy.ecotoxxplorer.ca
Low variance and low abundance features were filtered out and normalized using ecotoxxplorer.ca
The differentially expressed genes were identified using EdgeR, genes with adjusted p value <0.05 and log2 fold change value > 1 were considered significantly dysregulated. This was done in ecotoxxplorer.ca
Genome_build: Xenopus laevis genome (GenBank assembly accession: GCA_001663975.1)
Supplementary_files_format_and_content: csv - results of differentially expressed genes analysis using HiSat2
Supplementary_files_format_and_content: tabular - results of raw counts using Kallisto
|
| |
|
| Submission date |
Dec 03, 2021 |
| Last update date |
Aug 31, 2024 |
| Contact name |
Natacha Hogan |
| E-mail(s) |
natacha.hogan@usask.ca
|
| Organization name |
University of Saskatchewan
|
| Street address |
51 Campus Drive
|
| City |
Saskatoon |
| State/province |
SK |
| ZIP/Postal code |
S7N 5A8 |
| Country |
Canada |
| |
|
| Platform ID |
GPL28901 |
| Series (1) |
| GSE190080 |
Gene expression profiling and developmental outcomes reveals early toxicological mechanisms of lead effects in an early-life stage amphibian, Xenopus laevis |
|
| Relations |
| BioSample |
SAMN23577170 |
| SRA |
SRX13298289 |
