SummaryRMgm-5231
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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) : 35920043 |
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 | RMgm-1026 |
Other information parent line | A drug-selectable marker free reporter line expressing GFP under the constitutive hsp70 promoter |
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The mutant parasite was generated by | |
Name PI/Researcher | Fréville A, Khalife J |
Name Group/Department | Center for Infection and Immunity of Lille |
Name Institute | Univ. Lille, CNRS, Inserm, CHU Lille, Institut Pasteur de Lille |
City | Lille |
Country | France |
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Name of the mutant parasite | |
RMgm number | RMgm-5231 |
Principal name | PbLRR1KO |
Alternative name | |
Standardized name | |
Is the mutant parasite cloned after genetic modification | Yes |
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Phenotype | |
Asexual blood stage | Not different from wild type |
Gametocyte/Gamete | Not different from wild type |
Fertilization and ookinete | Not different from wild type |
Oocyst | A significant decrease (approx. 70%) in the number of oocysts at day 9 post-infection and oocysts at this stage of development were smaller in size. |
Sporozoite | Despite the overall smaller oocyst numbers and sizes in the PbLRR1KO lines, a substantial proportion of oocysts completed development to sporozoites and no significant effects on salivary gland colonization were observed. Sporozoites were infective in both in vitro hepatocyte cultures and in mice, albeit with a slightly longer prepatent period. |
Liver stage | Sporozoites were infective in both in vitro hepatocyte cultures and in mice, albeit with a slightly longer prepatent period. |
Additional remarks phenotype | Mutant/mutation Phenotype
A significant decrease (approx. 70%) in the number of oocysts at day 9 post-infection and oocysts at this stage of development were smaller in size.
Despite the overall smaller oocyst numbers and sizes in the PbLRR1KO lines, a substantial proportion of oocysts completed development to sporozoites and no significant effects on salivary gland colonization were observed. Sporozoites were infective in both in vitro hepatocyte cultures and in mice, albeit with a slightly longer prepatent period. Additional information Evidence is presented for an interaction of PbLRR1 to PbPP1c via 3 binding sites Other mutants |
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Details of the target gene | |||||||||||||||||||||||||
Gene Model of Rodent Parasite | PBANKA_0516600 | ||||||||||||||||||||||||
Gene Model P. falciparum ortholog | PF3D7_1032800 | ||||||||||||||||||||||||
Gene product | leucine-rich repeat protein | ||||||||||||||||||||||||
Gene product: Alternative name | PbLRR1 | ||||||||||||||||||||||||
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Details of the genetic modification | |||||||||||||||||||||||||
Inducable system used | No | ||||||||||||||||||||||||
Additional remarks inducable system | |||||||||||||||||||||||||
Type of plasmid/construct used | (Linear) plasmid double cross-over | ||||||||||||||||||||||||
PlasmoGEM (Sanger) construct/vector used | Yes | ||||||||||||||||||||||||
Name of PlasmoGEM construct/vector | PbGEM-232294 | ||||||||||||||||||||||||
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 | No | ||||||||||||||||||||||||
Additional remarks genetic modification | PbLRR1 KO lines were generated by double homologous recombination of a NotI-linearized PlasmoGEM vector. (PbGEM-232294, Wellcome Sanger Institute) transfected into PbGFP ANKA parasites. | ||||||||||||||||||||||||
Additional remarks selection procedure | |||||||||||||||||||||||||
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 | (Linear) 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 | No selectable marker | ||||||||||||||||||
Promoter of the selectable marker | eef1a | ||||||||||||||||||
Selection (positive) procedure | pyrimethamine | ||||||||||||||||||
Selection (negative) procedure | 5-fluorocytosine (5-FC) | ||||||||||||||||||
Additional remarks genetic modification | 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 | ||||||||||||||||||
Additional remarks selection procedure | |||||||||||||||||||
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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_0306000 | ||||||||||||||||||
Gene product | 6-cysteine protein | ||||||||||||||||||
Gene product: Alternative name | P230p | ||||||||||||||||||
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