RMgmDB - Rodent Malaria genetically modified Parasites


Malaria parasiteP. berghei
DisruptedGene model (rodent): PBANKA_0305900; Gene model (P.falciparum): PF3D7_0208800; Gene product: conserved Plasmodium protein, unknown function (pb22)
Phenotype Gametocyte/Gamete; Fertilization and ookinete; Oocyst;
Last modified: 27 November 2020, 17:40
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) : 33222390
MR4 number
Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. berghei
Parent strain/lineP. berghei ANKA
Name parent line/clone P. berghei ANKA 2.34
Other information parent lineP. berghei ANKA 2.34 is a cloned, gametocyte producer line of the ANKA strain (PubMed: PMID: 15137943).
The mutant parasite was generated by
Name PI/ResearcherLiu F, Cao Y
Name Group/DepartmentDepartment of Immunology, College of Basic Medical Sciences
Name InstituteChina Medical University
CityShenyang, Liaoning
Name of the mutant parasite
RMgm numberRMgm-4923
Principal namePb22-KO
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Asexual blood stageNot different from wild type
Gametocyte/GameteNormal numbers of mature male and female gametocytes are produced. Pb22-KO male gametocytes showed a ~89% reduction in the exflagellation centers formed. Normal formation (and egress) of female gametes.
Fertilization and ookineteStrongly reduced ookinete formation
OocystNo oocyst formation
SporozoiteNot tested
Liver stageNot tested
Additional remarks phenotype

The mutant lacks expression of Pb22

Protein (function)
PBANKA_0305900 (Pb22), which is predicted to encode a sexual-stage protein of 218 amino acids (aa) with a molecular weight of 22 kDa. Pb22 has a putative signal peptide (aa 1–18), suggesting that it might be secreted. Pb22 did not contain any domains of known functions, while the SMART program (http://smart.embl-heidelberg.de/) detected two low-complexity regions spanning aa 63–75 and aa 166–185, which contain compositionally biased amino acids. P22 is highly conserved among Plasmodium species. BLAST search also identified a homolog of this protein in Hepatocystis, which is considered to be descended from Plasmodium with the loss of blood schizogony.

Normal numbers of mature male and female gametocytes are produced. Pb22-KO male gametocytes showed a ~89% reduction in the exflagellation centers formed. Normal formation (and egress) of female gametes. Strongly reduced ookinete formation. No oocyst formation.
The major defects in the pb22-KO line appeared to be at the egress step since 78.9% of the microgametocytes in the pb22-KO line were in the stage of aborted exflagellation with the RBC membrane retained. For microgametocytes with aborted exflagellation, 30% failed to assemble the axonemes and 48.9% were egress-defective. 
Cross-fertilization experiments showed that the pb22-KO line was defective only in male fertility wheras female fertility was not affected.

Additional information
Immunofluorescence analyses showed the following: In the erythrocytic stages, Pb22 expression was restricted to the parasites in schizonts and gametocytes. Abundant expression of the Pb22 protein in both male and female gametocytes. In the exflagellating male gametes, the protein was observed on the residual body as well as the flagella. Evidence presented for surface location in gametes and ookinetes. 

Other mutants

  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_0305900
Gene Model P. falciparum ortholog PF3D7_0208800
Gene productconserved Plasmodium protein, unknown function
Gene product: Alternative namepb22
Details of the genetic modification
Inducable system usedNo
Additional remarks inducable system
Type of plasmid/construct used(Linear) plasmid double cross-over
PlasmoGEM (Sanger) construct/vector usedNo
Modified PlasmoGEM construct/vector usedNo
Plasmid/construct map
Plasmid/construct sequence
Restriction sites to linearize plasmid
Partial or complete disruption of the geneComplete
Additional remarks partial/complete disruption
Selectable marker used to select the mutant parasitehdhfr/yfcu
Promoter of the selectable markereef1a
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modificationPurified and digested PCR fragments (the 5’ and 3’ recombination fragments) were cloned into the PL0034 vector
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