SummaryRMgm-4996
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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) : 33762339 |
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 | Balestra AC, Brochet M |
Name Group/Department | Department of Microbiology and Molecular Medicine, Faculty of Medicine |
Name Institute | University of Geneva |
City | Geneva |
Country | Switzerland |
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Name of the mutant parasite | |
RMgm number | RMgm-4996 |
Principal name | P(clag)ICM1 |
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 | RT-PCR analysis confirmed a two-fold reduction in pbicm1 expression in purified P(clag)ICM1 gametocytes. Although P(clag)ICM1 gametocytes were microscopically indistinguishable from their wild-type (WT) counterparts, they did not form active exflagellation centers upon activation. PbICM1 down-regulation led to significant decreases in axoneme formation, egress from the host RBC, and DNA replication, indicating an early requirement for PbICM1 in male gametogenesis. A strongly attenuated calcium mobilization was consistently observed in P(clag)ICM1 |
Fertilization and ookinete | Not tested |
Oocyst | Not tested |
Sporozoite | Not tested |
Liver stage | Not tested |
Additional remarks phenotype | Mutant/mutation In the paper evidence is presented that: Analysis of a mutant expressing HA-tagged ICM1 (RMgm-4993) showed the following: Other mutants |
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Details of the target gene | |||||||||||||||||||||||||||
Gene Model of Rodent Parasite | PBANKA_1446100 | ||||||||||||||||||||||||||
Gene Model P. falciparum ortholog | Pf3D7_ 1231400 | ||||||||||||||||||||||||||
Gene product | membrane protein ICM1 | ||||||||||||||||||||||||||
Gene product: Alternative name | ICM1 | ||||||||||||||||||||||||||
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Details of the genetic modification | |||||||||||||||||||||||||||
Short description of the mutation | 'promoter swap mutant': the promoter of icm1 replaced with the clag9 promoter (PBANKA_140060). | ||||||||||||||||||||||||||
Inducable system used | No | ||||||||||||||||||||||||||
Short description of the conditional mutagenesis | Not available | ||||||||||||||||||||||||||
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 | hdhfr | ||||||||||||||||||||||||||
Promoter of the selectable marker | pbdhfr | ||||||||||||||||||||||||||
Selection (positive) procedure | pyrimethamine | ||||||||||||||||||||||||||
Selection (negative) procedure | No | ||||||||||||||||||||||||||
Additional remarks genetic modification | To generate the P(ama1)ICM1 line, the plasmid pOB116-ama1 was used (RMgm-773). The first 1000 bp from PbICM1 gene was amplified from gDNA using the primers 453 and 458. The PCR product was digested with Xho I and Not I and ligated into the pOB116-ama1 plasmid previously digested with the same enzymes, resulting in the plasmid pAB025. The last 750 bp of PbICM1 5′UTR were amplified using primers 459 and 464, and the resulting fragment was digested with Hind III and Pst I and ligated into pAB026 previously digested with the same enzymes. The same strategy was used to generate the line Pclag9ICM1, with the plasmid pOB116-clag9 resulting in plasmid pAB027. Both plasmids were digested with the enzymes Eco RV and Hind III to be transfected in P. berghei. | ||||||||||||||||||||||||||
Additional remarks selection procedure | |||||||||||||||||||||||||||
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