RMgmDB - Rodent Malaria genetically modified Parasites


Malaria parasiteP. berghei
DisruptedGene model (rodent): PBANKA_1134900; Gene model (P.falciparum): PF3D7_1358600; Gene product: zinc finger protein, putative (Pb103)
Phenotype Fertilization and ookinete; Oocyst; Sporozoite;
Last modified: 14 January 2022, 14:05
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) : 34959491
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/ResearcherHirai M, Mita T
Name Group/DepartmentDepartment of Tropical Medicine and Parasitology, Faculty of Medicine
Name InstituteJuntendo University
Name of the mutant parasite
RMgm numberRMgm-5142
Principal namePb103(-)
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Asexual blood stageNot different from wild type
Gametocyte/GameteNot different from wild type
Fertilization and ookineteAbsence of ookinete formation (due to abortion of zygote development).
OocystAbsence of ookinete and oocyst formation
SporozoiteAbsence of oocyst and sporozoite formation
Liver stageNot tested
Additional remarks phenotype

The mutant lacks expression of Pb103

Protein (function)
PBANKA_1134900 encodes 870 amino acids with a molecular weight of approximately 103 kDa, and designated it Pb103. InterProScan analysis revealed that Pb103 possesses two CCCH-type (zinc finger) ZF domains. Pb103 is highly conserved among rodent malaria parasites. While orthologous genes are  detected in human malaria parasites with low identity at the entire sequence level, two ZF domains are highly conserved among rodent and human malaria parasites.

Normal, wild type gametocyte/gamete formation. Absence of ookinete formation (due to abortion of zygote development). Absence of oocyst/sporozoite formation.

Additional information
RNAseq data shows that Pb103 is exclusively expressed in female gametocytes. 
Evidence is presented that transcripts are translationally repressed in female gametocytes (RMgm-5144). Translation occurs after activation of gametocytes.

In vitro cross-fertilization experiments were performed between Pb103(-) and CDPK4(-) (male infertility) or Nek4(-) (female infertility) gametes. The self-fertilization of CDPK4(-) and Nek4(-) did not produce ookinetes, while CDPK4(-) x Nek4(-) cross produced ookinetes, which was equivalent to 35.1% of CDPK4(-) female gametes fertilized to Nek4(-) males. Cross-fertilization of Pb103(-) x CDPK4(-) produced ookinetes with slightly lower efficiency compared to CDPK4(-) x Nek4(-) crosses. On the other hand, the Pb103(-) x Nek4(-) cross failed to generate mature ookinetes. These results demonstrate that Pb103 is exclusively required in females. 

To identify the domains critical for zygote/ookinete development, in this study several transgenic parasites expressing partially deleted Pb103 (RMgm-5143) were generated and assayed for ookinete maturation. Deleting either of two zinc finger (ZF) domains (ZFs) but not the C-terminal region abolished zygote/ookinete development, highlighting the indispensable roles of ZFs in parasite sexual development, most likely via translational repression.

Other mutants

  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_1134900
Gene Model P. falciparum ortholog PF3D7_1358600
Gene productzinc finger protein, putative
Gene product: Alternative namePb103
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
Promoter of the selectable markerpbdhfr
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modificationTo generate a construct for Pb103 gene disruption, two PCR fragments covering 500 bp of the 50 and 30 UTRs of the Pb103 gene were amplified by two pairs of primers, Pb03(-)- F1/Pb03(-)-R1 and Pb03(-)- F2/Pb03(-)-R2, respectively, and P. berghei genomic DNA as a template. Then, each PCR product was conjugated to either end of PstI/HindIII digested pL0006 (MRA-775; BEI Resources.) by fusion PCR using primer Pb03(-)-F1/Pb03(-)-R2
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