Back to search resultsSummaryRMgm-798
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Successful modification | The parasite was generated by the genetic modification |
The mutant contains the following genetic modification(s) | Gene mutation |
Reference (PubMed-PMID number) |
Reference 1 (PMID number) : 23245327 |
MR4 number | |
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Parent parasite used to introduce the genetic modification | |
Rodent Malaria Parasite | P. berghei |
Parent strain/line | P. berghei ANKA |
Name parent line/clone | P. berghei ANKA 2.34 |
Other information parent line | P. berghei ANKA 2.34 is a cloned, gametocyte producer line of the ANKA strain (PubMed: PMID: 15137943). |
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The mutant parasite was generated by | |
Name PI/Researcher | Pino, P.; Billker, O.; Soldati-Favre, D. |
Name Group/Department | Department of Microbiology and Molecular Medicine |
Name Institute | University of Geneva |
City | Geneva |
Country | Switzerland |
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Name of the mutant parasite | |
RMgm number | RMgm-798 |
Principal name | nmt-iKO |
Alternative name | |
Standardized name | |
Is the mutant parasite cloned after genetic modification | Yes |
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Phenotype | |
Asexual blood stage | Strongly reduced growth (parasitemia) in mice treated with ATc (see 'Additional information'). |
Gametocyte/Gamete | Not tested |
Fertilization and ookinete | Not tested |
Oocyst | Not tested |
Sporozoite | Not tested |
Liver stage | Not tested |
Additional remarks phenotype | Mutant/mutation Transctivator TRAD4 was made by a gene fusion between TetRep and an AD (activating domains/transactivating regions) of the ApiAP2 protein PFF0200c (= PfSIP2; fragment PfSIP2_7.3). |
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Details of the target gene | |||||||||||||||||||||||||||
Gene Model of Rodent Parasite | PBANKA_1029800 | ||||||||||||||||||||||||||
Gene Model P. falciparum ortholog | PF3D7_1412800 | ||||||||||||||||||||||||||
Gene product | glycylpeptide N-tetradecanoyltransferase | peptide N-myristoyltransferase | ||||||||||||||||||||||||||
Gene product: Alternative name | NMT | ||||||||||||||||||||||||||
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Details of the genetic modification | |||||||||||||||||||||||||||
Short description of the mutation | The nmt promoter modified to contain a tet-inducible promoter and TRAD4 activating domain | ||||||||||||||||||||||||||
Inducable system used | TET-based (TRAD4) | ||||||||||||||||||||||||||
Short description of the conditional mutagenesis | Downregulation of nmt expression by addition of the tetracycline derivative anhydrotetracycline(ATc) | ||||||||||||||||||||||||||
Additional remarks inducable system |
Conditional expression system for stage-specific, tetracycline-dependent (ATc) gene regulation. For in vivo treatment of mice, ATc (SIGMA) was dissolved in water + 5% sucrose at a concentration of 0.2 mg/ml. The drinking water bottle was wrapped in aluminum foil to prevent precipitation of ATc due to light and the solution changed every 48 hr. For in vitro treatments, ATc was diluted in culture medium at 1 mg/ml as previously described (Meissner et al., 2005). | ||||||||||||||||||||||||||
Type of plasmid/construct | Plasmid double cross-over | ||||||||||||||||||||||||||
PlasmoGEM (Sanger) construct/vector used | No | ||||||||||||||||||||||||||
Modified PlasmoGEM construct/vector used | No | ||||||||||||||||||||||||||
Plasmid/construct map | |||||||||||||||||||||||||||
Plasmid/construct sequence | |||||||||||||||||||||||||||
Restriction sites to linearize plasmid | |||||||||||||||||||||||||||
Selectable marker used to select the mutant parasite | hdhfr | ||||||||||||||||||||||||||
Promoter of the selectable marker | eef1a | ||||||||||||||||||||||||||
Selection (positive) procedure | pyrimethamine | ||||||||||||||||||||||||||
Selection (negative) procedure | No | ||||||||||||||||||||||||||
Additional remarks genetic modification | For tetracyclin (ATc)-dependent gene regulation the nmt locus was mutated as follows: A double crossing over recombination strategy was used to position TRAD4 downstream of the endogenous P. berghei nmt promoter, while the coding sequence of prf was brought under the control of the inducible promoter containing the tet operator. Simultaneously, two HA epitope tags were placed at the N terminus of the resulting inducible copy of nmt. Transctivator TRAD4 was made by a gene fusion between TetRep and an AD (activating domains/transactivating regions) of the ApiAP2 protein PFF0200c (= PfSIP2; fragment PfSIP2_7.3). | ||||||||||||||||||||||||||
Additional remarks selection procedure | |||||||||||||||||||||||||||
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