RMgmDB - Rodent Malaria genetically modified Parasites


Malaria parasiteP. berghei
TaggedGene model (rodent): PBANKA_1037800; Gene model (P.falciparum): PF3D7_1404300; Gene product: secreted ookinete adhesive protein, putative (SOAP)
Name tag: APEX2 + cmyc
Phenotype Fertilization and ookinete;
Last modified: 28 October 2020, 15:50
Successful modificationThe parasite was generated by the genetic modification
The mutant contains the following genetic modification(s) Gene tagging
Reference (PubMed-PMID number) Not published (yet)
MR4 number
Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. berghei
Parent strain/lineP. berghei ANKA
Name parent line/clone Not applicable
Other information parent line
The mutant parasite was generated by
Name PI/ResearcherKehrer J, frischknecht F
Name Group/DepartmentCenter for Infectious Diseases, Integrative Parasitology
Name InstituteHeidelberg University Medical School
Name of the mutant parasite
RMgm numberRMgm-4884
Principal namesoap::APEX2
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationNo
Asexual blood stageNot tested
Gametocyte/GameteNot tested
Fertilization and ookinetesee below
OocystNot tested
SporozoiteNot tested
Liver stageNot tested
Additional remarks phenotype

The mutant expresses a APEX2-cmyc tagged version of SOAP
Published in bioRxiv preprint doi: https://doi.org/10.1101/2020.09.29.318857.

Protein (function)

Additional information
To identify micronemal proteins of ookinetes we used the microneme-resident protein SOAP (Secreted Ookinete Adhesive Protein, PBANKA_1113400) as bait for proximity labeling. As ookinetes are fully formed within 21 h, but proximity based biotinylation with BirA* may require more than 24 h of incubation, we turned to the faster-acting ascorbate peroxidase APEX2 (Lam et al., 2014). SOAP is a small protein of 166 amino acids with a molecular weight of around 21 kDa and important for ookinete to oocyst transition (Dessens et al., 2003). In analogy to our previous approach (Kehrer, Frischknecht, et al., 2016) we generated a parasite line expressing SOAP C-terminally fused to APEX2 and a 3x myc tag. Fully developed and purified SOAP::APEX2 and wild type control ookinetes were incubated with biotin-phenol for 30 min at room temperature. The cells were then split into two separate populations and biotinylation was initiated in one vial for 1 min using H2O2 while the second vial served as control. Samples before and after enrichment of biotinylated proteins with streptavidin coated beads were tested for successful labeling of proteins by western blot and used for proteomic analyses. 

Other mutants

  Tagged: Mutant parasite with a tagged gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_1037800
Gene Model P. falciparum ortholog PF3D7_1404300
Gene productsecreted ookinete adhesive protein, putative
Gene product: Alternative nameSOAP
Details of the genetic modification
Name of the tagAPEX2 + cmyc
Details of taggingC-terminal
Additional remarks: tagging
Commercial source of tag-antibodies
Type of plasmid/construct(Linear) plasmid single 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 modificationsoap::APEX2: The APEX2::myc sequence flanked with the restriction sites BamHI and XbaI was ordered from geneart (Regensburg, Germany). It was ligated into the previously described BioID plasmid (Kehrer, Frischknecht, et al., 2016) replacing the bira*-sequence. The resulting plasmid pL8 was linearized with HpaI prior transfection for single crossover integration
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