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Status |
Public on Dec 12, 2022 |
Title |
wildtype, biological rep 5 |
Sample type |
SRA |
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Source name |
whole organism
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Organism |
Caenorhabditis elegans |
Characteristics |
tissue: whole organism cell line: AM141 cell type: whole organism genotype: 40Q::YFP treatment: 20 ºC, young adult
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Treatment protocol |
We sequenced 4 different genotypes, with 6 biological replicates per genotype.
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Growth protocol |
Synchronized L1 animals were cultured in NGM plates at 20 °C until they reached the young adult stage.
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Extracted molecule |
total RNA |
Extraction protocol |
Yound adults were collected in M9 buffer, frozen, thawed and mechanically lysed with 200 mg of glass beads (Sigma-Aldrich-Merck) for 30 sec using a Fastprep shaker apparatus (Thermofisher Scientific, FP120 model) . tRNA was extracted using NZY total RNA isolation kit, Nzytech. Supernatant was recovered from lysate samples by centrifuging extracts at maximum speed for 1 min. The purified RNA was treated with DNAse, quantified and sent for sequencing by Novogene (Cambridge, UK). Library preparation was performed at Novogene facilities (Cambridge, UK). mRNA was enriched using oligo(dT) beads and then fragmented randomly in fragmentation buffer, followed by cDNA synthesis using random hexamers and reverse transcriptase. After first-strand synthesis, a custom second-strand synthesis buffer (Illumina) was added with dNTPs, RNase H and Escherichia coli polymerase I to generate the second strand by nick- translation. To build the final cDNA library, a round of purification, terminal repair, A-tailing, ligation of sequencing adapters, size selection and PCR enrichment were performed. Library concentration was first quantified using a Qubit 2.0 fluorometer (Life Technologies). Insert size was checked on an Agilent 2100 and quantified using quantitative PCR (Q-PCR).
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Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
Illumina NovaSeq 6000 |
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Description |
A5_WT genes.readcount.annot.xls genes.FPKM.annot.xls nhr_1_vs_WT.genes.DEA.xls unc_1_vs_WT.genes.DEA.xls U_N_vs_WT.genes.DEA.xls
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Data processing |
Data filtering: Raw reads are filtered to remove reads with adapter contamination or reads with low quality. Only clean reads were used in the downstream analyses. The filtering process is as follows: (1) Discard reads with adaptor contamination. (2) Discard reads when uncertain nucleotides constitute more than 10% of either read (N > 10%). (3) Discard reads when low quality nucleotides (base quality less than 20) constitute more than 50% of the read. RNA-seq Adapter sequences (Oligonucleotide sequences of adapters from TruSeqTM RNA and DNA Sample Prep Kits): RNA 5' Adapter (RA5): 5'-AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATCT-3' RNA 3' Adapter (RA3): 5'-GATCGGAAGAGCACACGTCTGAACTCCAGTCAC(6-nucleotide index)ATCTCGTATGCCGTCTTCTGCTTG-3' Mapping: HISAT2 v2.0.5 with default parameters Quantification: HTSeq v0.6.1 with -m union option. Differential expression analysis: DESeq2 v1.24.0 Assembly: WBCel235 Supplementary files format and content: .bam file for each sample (24 total) Supplementary files format and content: .xls file with raw counts for each sample (1 column per sample) Supplementary files format and content: .xls file with FPKM counts for each sample (1 column per sample) Supplementary files format and content: .xls file from differential expression analysis for each pair of experimental groups (10 files total)
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Submission date |
Dec 11, 2022 |
Last update date |
Dec 12, 2022 |
Contact name |
Carlos Mora-Martínez |
E-mail(s) |
carmoma9@gmail.com
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Organization name |
Universidad Internacional de Valencia
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Street address |
Emilio Lluch 4Dup 16
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City |
Valencia |
State/province |
Comunidad Valenciana |
ZIP/Postal code |
46014 |
Country |
Spain |
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Platform ID |
GPL26672 |
Series (1) |
GSE220662 |
Changes in lipid metabolism driven by steroid signalling modulate proteostasis in C. elegans |
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Relations |
BioSample |
SAMN32153379 |
SRA |
SRX18638716 |
Supplementary data files not provided |
SRA Run Selector |
Raw data are available in SRA |
Processed data are available on Series record |
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