|
| Status |
Public on Mar 04, 2020 |
| Title |
N2 rep2 |
| Sample type |
SRA |
| |
|
| Source name |
Whole animal
|
| Organism |
Caenorhabditis elegans |
| Characteristics |
developmental stage: L4
strain: N2
genotype: wild-type
|
| Treatment protocol |
N/A
|
| Growth protocol |
Gravid wild-type and olrn-1(ums9) animals treated with 6% bleach and 5M NaOH to isolate eggs. Eggs from olrn-1(ums9) animals were harvested 18 hours before N2 due to the strain having a developmental delay. Animals were dropped to NGM plates seeded with OP50 and incubated at 20 degrees celcius untill the L4 stage when they were harvested.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated using TriReagent (Molecular Research Center, Inc.) and purified with RNeasy columns (Qiagen)
Oligo (dT) magnetic beads are used to select mRNA with polyA tail. Fragment the target RNA and reverse transcription to double-strand cDNA (dscDNA) by N6 random primer. End repair the dscDNA with phosphate at 5' end and stickiness 'A' at 3' end, then ligate and adaptor with stickiness 'T' at 3' end to the dscDNA. Two specific primers were used to amplify the ligation product. Denature the PCR product by heat and the single strand DNA is cyclized by splint oligo and DNA ligase.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
BGISEQ-500 |
| |
|
| Data processing |
The quality of raw sequencing data was evaluated by FastQC (Version 0.11.8) and multiqc (Version 1.7). Low-quality reads were trimmed using Trimmomatic (version 0.36). The trimmed reads were mapped to the C.elegans reference genome (WS220/ce11 on UCSC genome website) using HISAT2 (version 2.1.0). Then we use samtools (Version 1.3.1) to convert SAM files to sorted BAM files. GTF (WS220/ce11) annotation file was downloaded from UCSC genome website. The assembled gft file will be generated for each sample by using stringtie (Version 1.3.4). Stringtie will compare each sample against merged assembly and estimate it transcript abundance. Finally, we used stringtie to generate count table for Ballgown analysis. We use Ballgown package from the Bioconductor (Version 3.8) software suite and run custom R script in R console (R Version is 3.5) to analyze the gene differential expression, including: data visualization and inspection, statistical tests for differential expression, and multiple test correction. Differential gene expression was assessed on the multiple samples with the following parameters: fold change (FC) > 2 fold, adjusted P value < 0.05 and RPKM>1.
Genome_build: WS220/ce11
Supplementary_files_format_and_content: excel files include FPKM values for each sequenced sample
|
| |
|
| Submission date |
May 09, 2019 |
| Last update date |
Mar 04, 2020 |
| Contact name |
Read Pukkila-Worley |
| E-mail(s) |
read.pukkila-worley@umassmed.edu
|
| Phone |
508-856-3596
|
| Organization name |
University of Massachusetts Medical School
|
| Department |
Medicine
|
| Lab |
Pukkila-Worley
|
| Street address |
364 Plantation Street
|
| City |
Worcester |
| State/province |
MA |
| ZIP/Postal code |
01605 |
| Country |
USA |
| |
|
| Platform ID |
GPL25145 |
| Series (1) |
| GSE130959 |
Innate immunity in the C. elegans intestine is programmed by a neuronal regulator of AWC olfactory neuron development |
|
| Relations |
| BioSample |
SAMN11609777 |
| SRA |
SRX5811672 |
