RMgmDB - Rodent Malaria genetically modified Parasites


Malaria parasiteP. berghei
TaggedGene model (rodent): PBANKA_1429300; Gene model (P.falciparum): PF3D7_1213500; Gene product: integral membrane protein GPR180, putative (GPR180)
Name tag: triple-HA
Phenotype Asexual bloodstage; Gametocyte/Gamete; Fertilization and ookinete;
Last modified: 29 April 2022, 12:46
Successful modificationThe parasite was generated by the genetic modification
The mutant contains the following genetic modification(s) Gene tagging
Reference (PubMed-PMID number) Reference 1 (PMID number) : 35404079
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/ResearcherWang PP, Zhu Z
Name Group/DepartmentDepartment of Immunology, College of Basic Medical Sciences
Name InstituteChina Medical University
Name of the mutant parasite
RMgm numberRMgm-5197
Principal namePbGPR180-HA
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Asexual blood stageExpression of GPR180 in schizonts (not in ring stages and trophozoites), gametocytes and ookinetes with highest expression in gametocytes.
Gametocyte/GameteExpression of GPR180 in schizonts, gametocytes and ookinetes with highest expression in gametocytes. Evidence is presented that gametocyte activation stimulates GPR180 redistribution from the cytoplasm to the plasma membrane.
Fertilization and ookineteExpression of GPR180 in schizonts, gametocytes and ookinetes with highest expression in gametocytes.
OocystNot tested
SporozoiteNot tested
Liver stageNot tested
Additional remarks phenotype

The mutant expresses C-terminal 3xHA-tagged GPR180

Protein (function)
Heptahelical serpentine receptors are the largest group of membrane receptors responsible for transducing extracellular signals to various downstream effectors. The serpentine receptors coupled to heterotrimeric guanine nucleotide-binding proteins belong to G-protein-coupled receptors (GPCRs) with a salient feature of seven transmembrane domains, each consisting of 25–35 residues. Although there is little conservation in amino acid (aa) sequences across the entire GPCR superfamily, they share similar structures, which are used to classify the GPCRs into six main classes (A – F). Rhodopsin-like Class A is the largest class, accounting for around 90% of GPCRs. Structurally, Rhodopsin-like GPCRs have a GpcrRhopsn4 domain, an eighth helix, and a palmitoylated cysteine at the C-terminal tail. Bioinformatic analysis identified a gpr180-like gene in all Plasmodium species, with the GPCR-like transmembrane domain located in the C terminus, as predicted using the HMMER program. The ~250 aa GPCR domain contains residues that are highly conserved in the Rhodopsin-like GPCR transmembrane domain, which classifies the GPR180 proteins as Class A (Rhodopsin-like) family members of GPCRs

Expression of GPR180 in schizonts, gametocytes and ookinetes with highest expression in gametocytes. Evidence is presented that gametocyte activation stimulates GPR180 redistribution from the cytoplasm to the plasma membrane.

In schizonts, male and female gametocytes, the PbGPR180 fluorescence displayed diffused and occasionally punctate staining in the cytoplasm of the parasites. However, more PbGPR180 fluorescence was associated with the plasma membrane in female gametes, whereas it was mainly detected at the residual body of exflagellating male gametocytes. In both zygote and ookinete, PbGPR180 protein was predominantly associated with the plasma membrane. Furthermore, it was detected in ookinetes under both permeabilizing and nonpermeabilizing conditions, suggesting surface localization. 

Analysis of a mutant lacking expression of GPR180 (see RMgm-5196) indicates a function during male and female gamete formation (normal (wild type) numbers of gametocytes are produced; male gamete formation ~2fold reduced (as determined by counting exflagellation); female gamete formation reduced by ~ 30%. Crossing experiments indicate a more severe defect in female gametes compared to males. Reduced ookinete formation).

Additional information

Other mutants

  Tagged: Mutant parasite with a tagged gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_1429300
Gene Model P. falciparum ortholog PF3D7_1213500
Gene productintegral membrane protein GPR180, putative
Gene product: Alternative nameGPR180
Details of the genetic modification
Name of the tagtriple-HA
Details of taggingC-terminal
Additional remarks: tagging
Commercial source of tag-antibodies
Type of plasmid/construct(Linear) PCR construct double cross-over
PlasmoGEM (Sanger) construct/vector usedNo
Modified PlasmoGEM construct/vector usedNo
Plasmid/construct map
Plasmid/construct sequence
Restriction sites to linearize plasmid
Selectable marker used to select the mutant parasitetgdhfr
Promoter of the selectable markereef1a
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modificationTo tag the endogenous pbgpr180 with a 3xHA tag, an 1140 bp (nucleotide positions [nt] 938–2077 bp) was amplified from the P. berghei genomic DNA (gDNA) and ligated in the ApaI and SacII sites of the pL0034 plasmid as the 5' homologous region (5R). Then, the 871 bp of the pbgpr180 3'-untranslated region (UTR) (nt 11–1871 bp) was amplified and inserted between XhoI and NotI as the 3' homologous region (3R) to yield the final plasmid pL0034-PbGPR180-3xHA.
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