|
| Status |
Public on Oct 18, 2022 |
| Title |
Dd2M343Lcrt_repA |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Dd2-Dd2 CRT_M343L
|
| Organism |
Plasmodium falciparum |
| Characteristics |
culture: in vitro
Stage: Trophozoite asexual blood stage
strain: Dd2
genetic modification: ZFN-edited
pfcrt allele: Dd2_M343L
ppq sensitivity: Resistant
experimental replicate: Rep1
|
| Treatment protocol |
No drug treatment was performed.
|
| Growth protocol |
Parasite lines were cultured in vitro in 0.5% Albumax-supplemented RPMI 1640-based P. falciparum culture medium. Two consecutive 5% D-sorbitol treatments were used to tightly synchronize the cultures and magnetic column purifications were performed to obtain highly-enriched trophozoites when needed.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Extraction of total RNA, cDNA synthesis and amplification of target DNA was carried out as described in Bozdech, Z., Mok, S., & Gupta, A. P. DNA microarray-based genome-wide analyses of Plasmodium parasites. Methods Mol. Biol. 923, 189-211 (2013).
|
| Label |
Cy5
|
| Label protocol |
4ug of cDNA was labelled with Cy5 as described in Bozdech, Z., Mok, S., & Gupta, A. P. DNA microarray-based genome-wide analyses of Plasmodium parasites. Methods Mol. Biol. 923, 189-211 (2013). Samples were hybridized against a common Cy3-labelled pool consisting of RNA from mixed 3D7 asexual stages.
|
| |
|
| Channel 2 |
| Source name |
Plasmodium falciparum reference RNA pool
|
| Organism |
Plasmodium falciparum |
| Characteristics |
culture: in vitro
Stage: Asexual blood stage
strain: Pf3D7
composition: Mixture of equal amounts of RNA harvested every 6 hours over the 48-hour parasite's intraerythrocytic life cycle.
|
| Treatment protocol |
No drug treatment was performed.
|
| Growth protocol |
Parasite lines were cultured in vitro in 0.5% Albumax-supplemented RPMI 1640-based P. falciparum culture medium. Two consecutive 5% D-sorbitol treatments were used to tightly synchronize the cultures and magnetic column purifications were performed to obtain highly-enriched trophozoites when needed.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Extraction of total RNA, cDNA synthesis and amplification of target DNA was carried out as described in Bozdech, Z., Mok, S., & Gupta, A. P. DNA microarray-based genome-wide analyses of Plasmodium parasites. Methods Mol. Biol. 923, 189-211 (2013).
|
| Label |
Cy3
|
| Label protocol |
4ug of cDNA was labelled with Cy5 as described in Bozdech, Z., Mok, S., & Gupta, A. P. DNA microarray-based genome-wide analyses of Plasmodium parasites. Methods Mol. Biol. 923, 189-211 (2013). Samples were hybridized against a common Cy3-labelled pool consisting of RNA from mixed 3D7 asexual stages.
|
| |
|
| |
| Hybridization protocol |
Painter, H. J., Altenhofen, L. M., Kafsack, B. F. C. & Llinás, M. Whole-genome analysis of Plasmodium spp. Utilizing a new agilent technologies DNA microarray platform. Methods Mol. Biol. 923, 213–219 (2013).
|
| Scan protocol |
Agilent G2600D Microarray Scanner. Images were quantified using Agilent Feature Extraction Software (version 11.5.1.1).
|
| Description |
Biological replicate 1 of 3. PPQ-resistant PfCRT-edited M343L Dd2 line, untreated trophozoites.
|
| Data processing |
Agilent Feature Extraction Software (v 11.5.1.1) was used for background subtraction.
|
| |
|
| Submission date |
Jun 06, 2022 |
| Last update date |
Oct 18, 2022 |
| Contact name |
Manuel Llinas |
| E-mail(s) |
manuel@psu.edu
|
| Phone |
8148673444
|
| Organization name |
Penn State University
|
| Department |
Biochemistry and Molecular Biology
|
| Lab |
Manuel Llinas Lab
|
| Street address |
Millennium Science Complex, Pollock Rd
|
| City |
University Park |
| State/province |
Pennsylvania |
| ZIP/Postal code |
16802 |
| Country |
USA |
| |
|
| Platform ID |
GPL15130 |
| Series (1) |
| GSE205515 |
Piperaquine-resistant PfCRT mutations differentially impact drug transport, hemoglobin catabolism and parasite physiology in Plasmodium falciparum asexual blood stages |
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