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Sample GSM5956115 Query DataSets for GSM5956115
Status Public on Sep 17, 2022
Title Transcriptome of P. falciparum 3D7 treated with 625 nM CQ for 6 hours. Bioreplicate 2
Sample type RNA
 
Channel 1
Source name In vitro culture, asexual blood stage, laboratory strain 3D7, treated with antimalarial
Organism Plasmodium falciparum
Characteristics strain: 3D7
treatmant: CQ 625 nM
Treatment protocol Parasite culture was divided into 10 well in two 6-well plates with each well containing 6 mL of culture at 3% parasitaemia and 2% hematocrit and treated with 4-fold dilution series of CQ (10 µM top concentration), MK4815 (20 µM top concentration), MFQ (10 µM top concentration), primaquine (50 µM), and an equivalent volume of DMSO at 0.1% concentration and incubated for 6 hours at 37°C with 5% CO2, 3% O2, and 92% N2. Following incubation, the samples were harvested and centrifuged at 2,500 rpm for 5 minutes. Supernatant was removed and 1 mL of trizol was added the the pellets
Growth protocol P. falciparum strain 3D7 and Dd2 strains was obtained from BEI Resources and cultured under standard conditions in RPMI1640 media. Prior to drug treatment, parasites were double-synchronized with 5% sorbitol solution to achieve a synchrony of +/- 6 hours and cultured under constant agitation
Extracted molecule total RNA
Extraction protocol Total RNA was extracted using Trizol following manufacturer's instructions and as described in Bozdech, Z., S. Mok & A. P. Gupta, (2013) DNA microarray-based genome-wide analyses of Plasmodium parasites. Methods in molecular biology 923: 189-211.
Label Cy5
Label protocol Reverse transcription and cDNA synthesis were done following modifed Smart-Seq2 protocol (Picelli, S., Faridani, O., Björklund, Å. et al. Full-length RNA-seq from single cells using Smart-seq2. Nat Protoc 9, 171–181 (2014)). Synthesis of aminoallyl-coupled cDNA and labeling of samples with Cy fluorophore (GE Healthcare) were carried out as described in Bozdech, Z., S. Mok & A. P. Gupta, (2013) DNA microarray-based genome-wide analyses of Plasmodium parasites. Methods in molecular biology 923: 189-211.
 
Channel 2
Source name In vitro culture, asexual blood stage, laboratory strain 3D7, mixture of equal amounts of RNA harvested every 6 hours over the 48-hour parasite's intraerythrocytic life cycle
Organism Plasmodium falciparum
Characteristics condition: RNA reference pool
Treatment protocol Parasite culture was divided into 10 well in two 6-well plates with each well containing 6 mL of culture at 3% parasitaemia and 2% hematocrit and treated with 4-fold dilution series of CQ (10 µM top concentration), MK4815 (20 µM top concentration), MFQ (10 µM top concentration), primaquine (50 µM), and an equivalent volume of DMSO at 0.1% concentration and incubated for 6 hours at 37°C with 5% CO2, 3% O2, and 92% N2. Following incubation, the samples were harvested and centrifuged at 2,500 rpm for 5 minutes. Supernatant was removed and 1 mL of trizol was added the the pellets
Growth protocol P. falciparum strain 3D7 and Dd2 strains was obtained from BEI Resources and cultured under standard conditions in RPMI1640 media. Prior to drug treatment, parasites were double-synchronized with 5% sorbitol solution to achieve a synchrony of +/- 6 hours and cultured under constant agitation
Extracted molecule total RNA
Extraction protocol Total RNA was extracted using Trizol following manufacturer's instructions and as described in Bozdech, Z., S. Mok & A. P. Gupta, (2013) DNA microarray-based genome-wide analyses of Plasmodium parasites. Methods in molecular biology 923: 189-211.
Label Cy3
Label protocol Reverse transcription and cDNA synthesis were done following modifed Smart-Seq2 protocol (Picelli, S., Faridani, O., Björklund, Å. et al. Full-length RNA-seq from single cells using Smart-seq2. Nat Protoc 9, 171–181 (2014)). Synthesis of aminoallyl-coupled cDNA and labeling of samples with Cy fluorophore (GE Healthcare) were carried out as described in Bozdech, Z., S. Mok & A. P. Gupta, (2013) DNA microarray-based genome-wide analyses of Plasmodium parasites. Methods in molecular biology 923: 189-211.
 
 
Hybridization protocol Equal amounts of labelled sample and reference cDNA were hybridized on the microarray chip using the Agilent hybridization system. Hybridizations were performed for 20 hours at 70°C in a rotating hybridization oven at 10rpm as described in Bozdech, Z., S. Mok & A. P. Gupta, (2013) DNA microarray-based genome-wide analyses of Plasmodium parasites. Methods in molecular biology 923:
Scan protocol Microarray scanning was done using PowerScanner (Tecan) at 10uM resolution and with autogain PMT adjustments for both channels. Data was acquired using GenePix Pro v6.0 software (Axon Instruments) as described in Mok, S., M. Imwong et al (2011) Artemisinin resistance in Plasmodium falciparum is associated with an altered temporal pattern of transcription. BMC Genomics 12: 391.
Data processing Features with flag>0 and with median foreground intensity greater than 1.5 fold median background intensity for either channel passed quality control and were accepted.
Local background correction using normexp, followed by lowess normalization across all features within array and quantile normalization between arrays were carried out to generate the normalized Log2 gene expression ratios.
Background corrected and normalized log2 transformed ratios (Cy5/Cy3) of the RNA sample/3D7 RNA reference pool.
 
Submission date Mar 16, 2022
Last update date Sep 18, 2022
Contact name Zbynek Bozdech
E-mail(s) zbozdech@ntu.edu.sg
Phone 63162925
Organization name Nanyang Technological University
Street address 50 Nanyang Avenue
City Singapore
State/province Singapore
ZIP/Postal code 639798
Country Singapore
 
Platform ID GPL18893
Series (1)
GSE198746 Dose Response Transcriptional Profiling of Plasmodium falciparum 3D7 and Dd2 strains treated with chloroquine (CQ), MK-4815, and mefloquine (MFQ)

Supplementary file Size Download File type/resource
GSM5956115_3D7_CQ_B2_625_nM.gpr.gz 1.3 Mb (ftp)(http) GPR
Processed data are available on Series record

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