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| Status |
Public on Feb 21, 2019 |
| Title |
Hb_pellet_polyp_1 |
| Sample type |
SRA |
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| Source name |
Input pellet
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| Organism |
Heligmosomoides bakeri |
| Characteristics |
tissue: Input pellet
agent: polyphosphatase
treatment protocol: Nematodes were disrupted in 700 µl of Qiazol (Qiagen) using mechanical disruption with 5 mm stainless steel beads (Qiagen) on a Tissue Lyser II (30hz for 2 min twice; Qiagen)
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| Growth protocol |
CBA x C57BL/6 F1 (CBF1) mice were infected with L3 infective-stage H. bakeri larvae by gavage and adult nematodes were collected from the small intestine 14 days post infection. The nematodes were washed and maintained in serum-free media in vitro.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was treated with RNA 5' Polyphosphatase (Epicenter) following manufacturer's instructions, before library preparation.
Total RNA was extracted using a miRNAeasy mini kit (Qiagen) following manufacturer's instructions. RNA was treated with Turbo DNA-free kit (Thermo Fisher) to remove residual DNA.
Libraries were prepared using the CleanTag small RNA library prep kit according to manufacturer’s instructions, using total RNA from adult worms (30 ng), and sucrose-gradient purified EVs (equivalent 1E10 EVs measured by Nanosight, Malvern). For all samples, 1:12 dilutions of both adapters were used with 18 amplification cycles (TriLink BioTechnologies). Fragments between 140-170bp were size selected and sequenced on an Illumina HiSeq high output v4 50bp SE in Edinburgh Genomics.
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| Library strategy |
ncRNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
All libraries were quality checked with FastQC (http://www.bioinformatics.babraham.ac.uk/projects/fastqc/), reaper v16-098 (https://www.ebi.ac.uk/~stijn/reaper/reaper.html) was used to remove the Illumina small RNA Adapter sequences, and PullSeq 1.0.2 (https://github.com/bcthomas/pullseq) to keep reads that were at least 16 nucleotides long.
Tally v16-098 (https://www.ebi.ac.uk/~stijn/reaper/tally.html) was used to collapse all reads to a single occurrence in a FASTA file, keeping the counts of each read in the header. These are provided as processed files.
The alignment component of ShortStack 3.8.3 was used, with parameters: --nostitch, --mismatches 2, --mmap u, --bowtie_m 500 and --ranmax 500.
All the sRNA-seq results for each genome were used to predict novel ncRNA producing regions or clusters. All libraries for each genome were combined as a single input, and ShortStack 3.8.3 was used with the following parameters: --pad 10, --mincov 10, --dicermin 18, --dicermax 32. All the clusters were then split at exon-intron boundaries.
In order to obtain a level of expression for each sRNA-producing cluster, mapped reads were counted using the findOverlaps function from GenomicRanges R package with parameters minoverlap=16 and ignore.strand=TRUE. Only reads between 20-25nt were counted. Count tables were produced for each genome, where each cluster is a row and each library a column. These are provided as supplementary files.
Genome_build: The reference genome assembly for H. bakeri genome was nHp_v2.0 from http://parasite.wormbase.org/Heligmosomoides_polygyrus_prjeb15396 and for C. elegans WBcel235 from http://parasite.wormbase.org/Caenorhabditis_elegans_prjna13758.
Supplementary_files_format_and_content: Contains all reads of at least 16 nucleotides collapsed to a unique occurrences and saved in FASTA format. Read count is included in the tab files.
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| Submission date |
Jul 16, 2018 |
| Last update date |
Jun 16, 2020 |
| Contact name |
Cei Abreu-Goodger |
| E-mail(s) |
cei.abreu@cinvestav.mx
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| Organization name |
Langebio - Cinvestav
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| Street address |
Km. 9.6 Libramiento Nte Carr Leon-Irapuato
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| City |
Irapuato |
| State/province |
Guanajuato |
| ZIP/Postal code |
36824 |
| Country |
Mexico |
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| Platform ID |
GPL25342 |
| Series (1) |
| GSE117169 |
An extracellular Argonaute protein mediates export of repeat-associated small RNAs into vesicles in parasitic nematodes |
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| Relations |
| Reanalyzed by |
GSM4618495 |
| BioSample |
SAMN09665763 |
| SRA |
SRX4394664 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3272810_Hb_pellet_polyp_1.fa.gz |
5.2 Mb |
(ftp)(http) |
FA |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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