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| Status |
Public on May 08, 2015 |
| Title |
Ex vivo2 |
| Sample type |
SRA |
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| Source name |
ex vivo incubation
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| Organism |
Glaesserella parasuis str. Nagasaki |
| Characteristics |
strain: Nagasaki
culture time: 24 h
genotype: wild type
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| Growth protocol |
24h chocolate agar plate culture at 37ºC
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| Extracted molecule |
total RNA |
| Extraction protocol |
Sequencing library was generated using an Ion Torrent RNA-Seq v2 kit (Life Technologies) and RNA was sequenced using an Ion Torrent PGM instrument (Life Technologies) with an Ion 316 chip (Life technologies) at the Centre for Research in Agricultural Genomics (CRAG, Campus de Bellaterra-UAB, Spain). As control, RNA from the Nagasaki strain grown overnight on chocolate agar plates was purified and sequenced in the same manner.
Bacteria were recovered by centrifugation and pellets were used for RNA extraction. An additional low speed centrifugation step was added to ex vivo and in vivo samples in order to eliminate mammalian cells. All bacterial samples were processed for RNA extraction as follows. Pellets were resuspended in 2% SDS 16 mM EDTA 10 mM TRIS (pH XX) and incubated for 5 min at 100 ºC. Afterwards, samples were processed by two hot acid phenol-chloroform extractions, followed by two chloroform/ isoamyl alcohol extractions. RNA was then precipitated with 0.6 volumes of isopropanol 0.1 volumes of 5M ammonium acetate and 1μL of glycogen. After centrifugation the pellet was washed with 70% ethanol, dried and resuspended in warmed RNase-free water. To ensure that contaminating bacterial DNA was eliminated from the samples, treatment with RNase-free DNase (Qiagen) was performed. In addition, ribosomal RNA was eliminated with the Ribo-Zero rRNA removal kit (Epicentre Biotechnologies, Madison, WI, USA) following manufacturer’s instructions. PCR reactions using primers specific for H. parasuis 16S rRNA gene were carried out to ensure that no bacterial DNA was left in the sample.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Ion Torrent PGM |
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| Description |
ABKM00000000.1
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| Data processing |
For both in vivo and ex vivo experiments, bioinformatic analysis was performed following the count-based differential expression method (Anders et al., 2013), with some modifications.
Genome_build: Haemophilus parasuis str. Nagasaki
Supplementary_files_format_and_content: Tab-delimited text files include counts for each gene.
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| Submission date |
May 08, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
BERNARDO BELLO ORTI |
| E-mail(s) |
[email protected]
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| Organization name |
CReSA
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| Department |
Bacteriology
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| Street address |
Bellaterra
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| City |
Barcelona |
| State/province |
Barcelona |
| ZIP/Postal code |
08193 |
| Country |
Spain |
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| Platform ID |
GPL19492 |
| Series (1) |
| GSE63851 |
Haemophilus parasuis gene expression the in lung |
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| Relations |
| BioSample |
SAMN03647090 |
| SRA |
SRX1022032 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1678900_exvivo2_F.counts.txt.gz |
9.8 Kb |
(ftp)(http) |
TXT |
| GSM1678900_exvivo2_R.counts.txt.gz |
8.3 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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