Back to search resultsSummaryRMgm-1030
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Successful modification | The parasite was generated by the genetic modification |
The mutant contains the following genetic modification(s) | Gene disruption, Introduction of a transgene |
Reference (PubMed-PMID number) |
Reference 1 (PMID number) : 24755823 |
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 cl15cy1 |
Other information parent line | A reference wild type clone from the ANKA strain of P. berghei (PubMed: PMID: 17406255). |
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The mutant parasite was generated by | |
Name PI/Researcher | Manzoni G; Silvie O |
Name Group/Department | INSERM |
Name Institute | INSERM |
City | Paris |
Country | France |
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Name of the mutant parasite | |
RMgm number | RMgm-1030 |
Principal name | Δslarp-GFP |
Alternative name | |
Standardized name | |
Is the mutant parasite cloned after genetic modification | Yes |
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Phenotype | |
Asexual blood stage | Not tested |
Gametocyte/Gamete | Not tested |
Fertilization and ookinete | Not tested |
Oocyst | Not tested |
Sporozoite | Not tested |
Liver stage | To analyse in more details the fate of Δslarp sporozoites, time-lapse fluorescence microscopy was used to image control Δp230p-GFP (RMgm-1026) and Δslarp-GFP parasites in HepG2 cultures from 7 to 24 hours postinfection. Transformation of elongated sporozoites into round forms was observed with both Δp230p-GFP and Δslarp-GFP. However, whilst Δp230p-GFP persisted in the culture and started growing over the observation period, Δslarp-GFP failed to develop and died, as evidence by disappearance of the GFP signal from infected cells. No detectable sign of death of infected host cells was observed by microscopy, suggesting that elimination of the Δslarp mutant parasites, unlike other liver stage-arresting mutants19, does not result from host cell apoptosis. |
Additional remarks phenotype | Mutant/mutation To analyse in more details the fate of Δslarp sporozoites, time-lapse fluorescence microscopy was used to image control Δp230p-GFP (RMgm-1026) and Δslarp-GFP parasites in HepG2 cultures from 7 to 24 hours postinfection. Transformation of elongated sporozoites into round forms was observed with both Δp230p-GFP and Δslarp-GFP. However, whilst Δp230p-GFP persisted in the culture and started growing over the observation period, Δslarp-GFP failed to develop and died, as evidence by disappearance of the GFP signal from infected cells. No detectable sign of death of infected host cells was observed by microscopy, suggesting that elimination of the Δslarp mutant parasites, unlike other liver stage-arresting mutants19, does not result from host cell apoptosis. Other mutants |
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Details of the target gene | |||||||||||||||||||||||||
Gene Model of Rodent Parasite | PBANKA_0902100 | ||||||||||||||||||||||||
Gene Model P. falciparum ortholog | PF3D7_1147000 | ||||||||||||||||||||||||
Gene product | sporozoite and liver stage asparagine-rich protein | sporozoite asparagine-rich protein | ||||||||||||||||||||||||
Gene product: Alternative name | SLARP; S22; SAP1 | ||||||||||||||||||||||||
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Details of the genetic modification | |||||||||||||||||||||||||
Inducable system used | No | ||||||||||||||||||||||||
Additional remarks inducable system | |||||||||||||||||||||||||
Type of plasmid/construct used | 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 | |||||||||||||||||||||||||
Partial or complete disruption of the gene | Complete | ||||||||||||||||||||||||
Additional remarks partial/complete disruption | |||||||||||||||||||||||||
Selectable marker used to select the mutant parasite | hdhfr/yfcu | ||||||||||||||||||||||||
Promoter of the selectable marker | eef1a | ||||||||||||||||||||||||
Selection (positive) procedure | pyrimethamine | ||||||||||||||||||||||||
Selection (negative) procedure | 5-fluorocytosine (5-FC) | ||||||||||||||||||||||||
Additional remarks genetic modification | The parasites have been generated and selected using the GOMO method of transfection (see RMgm-1026 for more details for this transfection method) | ||||||||||||||||||||||||
Additional remarks selection procedure | The parasites are selected by a combination of positive selection (pyrimethamine), negative selection (5-FC) and FACS sorting. 1) Transfected parasites are first selected in a mouse by pyrimethamine treatment 2) GFP+mCherry+ parasites are selected by FACS sorting and used to infect a mice 3) This mouse is treated with 5-FC to select for parasites that have the selectable marker removed 4) GFP+mCherry- and marker free parasites are selected by FAC sorting and used to infect a mouse | ||||||||||||||||||||||||
Primer information: Primers used for amplification of the target sequences
Primer information: Primers used for amplification of the target sequences
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Type and details of transgene | |||||||||||||||||||
Is the transgene Plasmodium derived | Transgene: not Plasmodium | ||||||||||||||||||
Transgene name | GFP | ||||||||||||||||||
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Details of the genetic modification | |||||||||||||||||||
Inducable system used | No | ||||||||||||||||||
Additional remarks inducable system | |||||||||||||||||||
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/yfcu | ||||||||||||||||||
Promoter of the selectable marker | eef1a | ||||||||||||||||||
Selection (positive) procedure | pyrimethamine | ||||||||||||||||||
Selection (negative) procedure | 5-fluorocytosine (5-FC) | ||||||||||||||||||
Additional remarks genetic modification | |||||||||||||||||||
Additional remarks selection procedure | The parasites are selected by a combination of positive selection (pyrimethamine), negative selection (5-FC) and FACS sorting. 1) Transfected parasites are first selected in a mouse by pyrimethamine treatment 2) GFP+mCherry+ parasites are selected by FACS sorting and used to infect a mice 3) This mouse is treated with 5-FC to select for parasites that have the selectable marker removed 4) GFP+mCherry- and marker free parasites are selected by FAC sorting and used to infect a mouse | ||||||||||||||||||
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Other details transgene | |||||||||||||||||||
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Promoter | |||||||||||||||||||
Gene Model of Parasite | PBANKA_0711900 | ||||||||||||||||||
Gene Model P. falciparum ortholog | PF3D7_0818900 | ||||||||||||||||||
Gene product | heat shock protein 70 | ||||||||||||||||||
Gene product: Alternative name | HSP70 | ||||||||||||||||||
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3'-UTR | |||||||||||||||||||
Gene Model of Parasite | PBANKA_0719300 | ||||||||||||||||||
Gene product | bifunctional dihydrofolate reductase-thymidylate synthase, putative | ||||||||||||||||||
Gene product: Alternative name | dhfr/ts | ||||||||||||||||||
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Insertion/Replacement locus | |||||||||||||||||||
Replacement / Insertion | Replacement locus | ||||||||||||||||||
Gene Model of Parasite | PBANKA_0902100 | ||||||||||||||||||
Gene product | sporozoite and liver stage asparagine-rich protein sporozoite asparagine-rich protein | ||||||||||||||||||
Gene product: Alternative name | SLARP; S22; SAP1 | ||||||||||||||||||
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