RMgmDB - Rodent Malaria genetically modified Parasites


Malaria parasiteP. berghei
TaggedGene model (rodent): PBANKA_0205600; Gene model (P.falciparum): PF3D7_0107800; Gene product: double-strand break repair protein MRE11 (MRE11)
Name tag: GFP
Phenotype Gametocyte/Gamete; Fertilization and ookinete; Oocyst; Sporozoite;
Last modified: 2 June 2021, 13:39
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) : 33287434
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/ResearcherGuttery DS, Tewari R
Name Group/DepartmentSchool of Life Sciences
Name InstituteUniversity of Nottingham
Name of the mutant parasite
RMgm numberRMgm-4877
Principal nameMRE11-GFP
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Asexual blood stageNot different from wild type
Gametocyte/GameteMRE11-GFP present in nucleus of female gametocytes (not in males)
Fertilization and ookineteFollowing gametocyte activation, female, but not male, gametocytes continued to express MRE11-GFP. After fertilization the protein remained associated with the nucleus throughout zygote/ookinete development
OocystIn oocysts the MRE11-GFP expression was diffuse
SporozoiteIn sporozoites MRE11-GFP was focused at a single point adjacent to the nuclear DNA
Liver stageNot tested
Additional remarks phenotype

The mutant expresses a C-terminal GFP-tagged version of MRE11.

Protein (function)
Double-strand breaks in the DNA resulting from innate DNA replication errors or exposure to radiation and chemical mutagens activate the DSB repair (DSBR) pathway, facilitating repair via two distinct mechanisms: “error-prone” non-homologous end-joining (NHEJ) and “error-free” homologous recombination (HR). In Plasmodium, HR appears to be the predominant DDR pathway, with homologues of many key players of mammalian HR. A key initiator of the HR pathway is meiotic recombination 11 protein (MRE11), which in mammals is part of the MRN/X complex that includes MRE11, RAD50 and NBS1 (Nijmegen breakage syndrome 1, a homologue of Saccharomyces cerevisae Xrs2). This complex has a crucial role in DDR during mitosis, promoting HR between sister chromatids to repair mutations arising during DNA replication. MRE11 is not essential in Trypanosoma brucei bloodstream forms, but its deletion results in impaired HR, reduced growth and increased sensitivity to DNA double-strand breaks. Numerous proteins potentially involved in HR have been identified in P. falciparum, although NBS1 is not encoded in the genome. In a recent study, a single mre11 orthologue containing 2 incomplete but catalytically active MPP_MRE11 domains was identified in P. falciparum (Gene ID: PF3D7_0107800), and complementation experiments confirmed DDR activity in response to DNA damage. A role in DDR was confirmed when P. falciparum mre11 (along with its MRN partner rad50) was shown to be significantly upregulated in response to treatment to induce DNA damage with the alkylating agent, methyl methanesulfonate (MMS).

Analyses of a mutant lacking MRE11 (RMgm-4876) showed a role of MRE11 in formation of mature oocysts. The Δmre11 oocysts had a fully formed wall but the cytoplasm exhibited evidence of degeneration, with vacuolization and organelle breakdown, while the nuclei showed marked swelling of the nuclear membranes. No salivary gland sporozoites are formed

Analyses of the mutant expressing a GFP-tagged MR11 showed:
MRE11-GFP was not present in asexual blood stage parasites, but was detected in the first sexual stages, more specifically in the nucleus of female but not male gametocytes. Following gametocyte activation, female, but not male, gametocytes continued to express the protein. After fertilization the protein remained associated with the nucleus throughout zygote/ookinete development; whereas in oocysts the expression was diffuse and in sporozoites it was focused at a single point adjacent to the nuclear DNA 

Additional information
Evidence is presented that: 
Mre11 mRNA is present in cells throughout the P. berghei life-cycle (except merozoites), based on single cell RNA-seq data, with the highest abundance in ookinetes/oocysts.


Other mutants

  Tagged: Mutant parasite with a tagged gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_0205600
Gene Model P. falciparum ortholog PF3D7_0107800
Gene productdouble-strand break repair protein MRE11
Gene product: Alternative nameMRE11
Details of the genetic modification
Name of the tagGFP
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 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