Summary

RMgm-1
Malaria parasiteP. berghei
Genotype
DisruptedGene model (rodent): PBANKA_0514900; Gene model (P.falciparum): PF3D7_1030900; Gene product: 28 kDa ookinete surface protein (Pbs21; P28)
Phenotype Fertilization and ookinete; Oocyst;
Last modified: 6 June 2009, 23:20
  *RMgm-1
Successful modificationThe parasite was generated by the genetic modification
The mutant contains the following genetic modification(s) Gene disruption
Reference (PubMed-PMID number) Reference 1 (PMID number) : 11483501
MR4 number MRA-442
Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. berghei
Parent strain/lineP. berghei ANKA
Name parent line/clone P. berghei ANKA cl15cy1
Other information parent lineA reference wild type clone from the ANKA strain of P. berghei (PubMed: PMID: 17406255).
The mutant parasite was generated by
Name PI/ResearcherC.J. Janse, A.P. Waters, A.M. Thomas
Name Group/DepartmentLeiden Malaria Research Group
Name InstituteLeiden University Medical Center
CityLeiden
CountryThe Netherlands
Name of the mutant parasite
RMgm numberRMgm-1
Principal name60cl3; 66cl1
Alternative name28Sko (Pbs21ko)
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Phenotype
Asexual blood stageNot different from wild type
Gametocyte/GameteNot different from wild type
Fertilization and ookineteSlight inhibition of ookinete development (slight reduction in ookinete numbers in in vitro cultures); ookinetes do not form the typical clusters of multiple ookinetes in vitro
OocystSlight inhibition of oocyst development (slight reduction in ookinete numbers in in Anopheles stephensi mosquitoes).
SporozoiteNot different from wild type
Liver stageNot different from wild type
Additional remarks phenotype

Mutant/mutation
The mutant lacks expression of the ookinete surface protein P28.

Protein (function)
P28 is one of two major surface proteins (P25 and P28) of the plasma membrane of ookinetes. The proteins are conserved between different Plasmodium species. The proteins are characterized by a secretory N-terminal signal sequence followed by three or four epidermal growth factor (EGF) domains and a glycosylphosphatidylinositol (GPI) anchor. Synthesis of both proteins begins between 0.5 and 2 h after the formation of the female gametes, and the proteins are still present in young oocysts. The paralogous genes encoding P25 and P28 are located next to each other in the genome in a head to tail organization.

Phenotype
The phenotype analyzes show only a minor effect on ookinete and oocyst development, indicating a redundancy in function of P25 and P28 (see also mutant RMgm-3 that lacks expression of both P25 and P28).

Additional information
Motility and entry into insect cells in vitro of ookinetes lacking expression of P28 are normal, but the number of ookinetes successfully transforming into oocysts in vitro was significantly reduced (Siden-Kiamos et al., J. Cell Science (2000) 113, 3419-26).

Other mutants
A. P. berghei mutant has been generated lacking P25 (RMgm-2).
A. P. berghei mutant has been generated lacking both P28 and P25 (RMgm-3).


  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_0514900
Gene Model P. falciparum ortholog PF3D7_1030900
Gene product28 kDa ookinete surface protein
Gene product: Alternative namePbs21; P28
Details of the genetic modification
Inducable system usedNo
Additional remarks inducable system
Type of plasmid/construct usedPlasmid double cross-over
PlasmoGEM (Sanger) construct/vector usedNo
Modified PlasmoGEM construct/vector usedNo
Plasmid/construct map
Plasmid/construct sequence
Restriction sites to linearize plasmid NheI, SpeI
Partial or complete disruption of the geneComplete
Additional remarks partial/complete disruption
Selectable marker used to select the mutant parasitetgdhfr
Promoter of the selectable markerpbdhfr
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modification
Additional remarks selection procedure
Primer information: Primers used for amplification of the target sequences  Click to view information
Primer information: Primers used for amplification of the target sequences  Click to hide information
Sequence Primer 1
Additional information primer 1
Sequence Primer 2
Additional information primer 2
Sequence Primer 3
Additional information primer 3
Sequence Primer 4
Additional information primer 4
Sequence Primer 5
Additional information primer 5
Sequence Primer 6
Additional information primer 6