|
| Status |
Public on Jun 10, 2020 |
| Title |
E5ind_rapa_40-45_rep2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
E5ind, rapa., 40-45 hpi
|
| Organism |
Plasmodium falciparum |
| Characteristics |
strain: E5ind
treatment: Rapamycin
|
| Treatment protocol |
Tightly synchronized E5ind cultures (0-5 h) were induced with 10 nM rapamycin (Rapa) for 1 h at 25-30 hpi or with DMSO as control (Dmso).
|
| Growth protocol |
Parasites were cultured in B+ erythrocytes at a 3 % haematocrit under standard culture conditions in RPMI-based media containing Albumax II without human serum, in a 5% CO2, 3% O2, balance N2 atmosphere. WR99210 selection was always maintained until the sychronization of the cultures. Tight synchronization (5 h age window) was achieved by Percoll purification followed by sorbitol treatment 5 h later.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was purified using the TRIzol method following the manufacturer intructions.
|
| Label |
Cy5
|
| Label protocol |
Test sample, Cy5; reference pool, Cy3. As previously described (Painter, H. et al., 2013. PMID: 22990780)
|
| |
|
| Channel 2 |
| Source name |
E5 wild-type and E5ind reference pool
|
| Organism |
Plasmodium falciparum |
| Characteristics |
strain: E5 and E5ind
time-point: 6 samples of synchronized cultures covering the whole asexual cycle.
|
| Treatment protocol |
Tightly synchronized E5ind cultures (0-5 h) were induced with 10 nM rapamycin (Rapa) for 1 h at 25-30 hpi or with DMSO as control (Dmso).
|
| Growth protocol |
Parasites were cultured in B+ erythrocytes at a 3 % haematocrit under standard culture conditions in RPMI-based media containing Albumax II without human serum, in a 5% CO2, 3% O2, balance N2 atmosphere. WR99210 selection was always maintained until the sychronization of the cultures. Tight synchronization (5 h age window) was achieved by Percoll purification followed by sorbitol treatment 5 h later.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was purified using the TRIzol method following the manufacturer intructions.
|
| Label |
Cy3
|
| Label protocol |
Test sample, Cy5; reference pool, Cy3. As previously described (Painter, H. et al., 2013. PMID: 22990780)
|
| |
|
| |
| Hybridization protocol |
Hybridization performed as previously described (Painter, H. et al., 2013. PMID: 22990780).
|
| Scan protocol |
Microarrays images were obtained using the Agilent G2505C Microarray Scanner.
|
| Data processing |
Initial data processing of microarray data, including normalization, was performed using the GE2_1105_Oct12 extraction protocol in Agilent Feature Extraction 11.5.1.1 software. The next steps of the analysis were performed using Bioconductor in an R environment (R version 3.5.3). For each individual microarray, we calculated Cy3 and Cy5 background signal as the median of the 100 lowest signal probes for each channel. Probes with both Cy3 and Cy5 signals below three times the array background were excluded. Gene level log2(Cy5/Cy3) values and statistical estimation of parasite age were computed as described (Rovira-Graells et al., 2012). Genes with missing values or genes that in all samples had expression values within the lowest 20th percentile of intensity (Cy5 channel) were excluded from downstream analysis, including identification of differentially expressed genes, because differences in genes expressed at near-background levels are of low confidence.
|
| |
|
| Submission date |
Jul 29, 2019 |
| Last update date |
Jun 11, 2020 |
| Contact name |
Oriol Llorà-Batlle |
| E-mail(s) |
oriol.llora@isglobal.org
|
| Phone |
+34932275400-4080
|
| Organization name |
ISGlobal
|
| Street address |
Carrer Rosselló 153, 1st floor
|
| City |
Barcelona |
| ZIP/Postal code |
08036 |
| Country |
Spain |
| |
|
| Platform ID |
GPL26985 |
| Series (2) |
| GSE135005 |
Conditional expression of PfAP2-G for controlled massive sexual conversion in Plasmodium falciparum [array] |
| GSE135006 |
Conditional expression of PfAP2-G for controlled massive sexual conversion in Plasmodium falciparum |
|
