RMgmDB - Rodent Malaria genetically modified Parasites

Summary

RMgm-5527
Malaria parasiteP. berghei
Genotype
TaggedGene model (rodent): PBANKA_0519900; Gene model (P.falciparum): PF3D7_1036900; Gene product: conserved Plasmodium protein, unknown function
Name tag: GFP
Phenotype Asexual bloodstage;
Last modified: 8 July 2024, 13:13
  *RMgm-5527
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) : 38195464
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/ResearcherNiikura M, Kobayashi F
Name Group/DepartmentDepartment of Environmental Science, School of Life and Environmental Science
Name InstituteAzabu University
CityKanagawa
CountryJapan
Name of the mutant parasite
RMgm numberRMgm-5527
Principal name0519900::GFP
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Phenotype
Asexual blood stageEvidence for localization in the cytoplasm of asexual blood stages.
Gametocyte/GameteNot tested
Fertilization and ookineteNot tested
OocystNot tested
SporozoiteNot tested
Liver stageNot tested
Additional remarks phenotype

Mutant/mutation
The mutant expresses a C-terminal GFP-tagged version of of PBANKA_0519900

Protein (function)
To investigate the proteins that interact with the NTF2 domain-containing protein PBANKA_1019700, protein, immunoprecipitation (IP) using anti-mCherry beads was performed and the proteins bound to PBANKA_1019700 were identified through mass spectrometry (MS) (see RMgm-5525). Cytoplasmic proteins, including ubiquitin-related proteins (ubiquitin carboxylterminal hydrolase, PBANKA_0210600; ubiquitin-like protein, PBANKA_0823000; HECT-type E3 ubiquitin ligase UT, PBANKA_0802300) and a VPS13 domain-containing protein (PBANKA_1359300) were detected through IP-MS of 1019700::mCherry. However, RNA-binding proteins, such as NAB2 and GBP2, were not detected. In addition, the conserved Plasmodium protein PBANKA_0519900 was detected

Phenotype
Analysis of the mutant expressing a C-terminal GFP-tagged version of PBANKA_1019700 showed evidence for localization in the cytoplasm of asexual blood stages.

Analysis of a mutant lacking expression of PBANKA_0519900 (RMgm-2526) showed the following: mice infected with Δ0519900 mutants showed lower levels of parasitaemia than mice infected with control parasites and their survival was prolonged compared to mice infected with control parasites

Additional information
To investigate the proteins that interact with the NTF2 domain-containing protein PBANKA_1019700, protein, immunoprecipitation (IP) using anti-mCherry beads was performed and the proteins bound to PBANKA_1019700 were identified through mass spectrometry (MS) (see RMgm-5525). Cytoplasmic proteins, including ubiquitin-related proteins (ubiquitin carboxylterminal hydrolase, PBANKA_0210600; ubiquitin-like protein, PBANKA_0823000; HECT-type E3 ubiquitin ligase UT, PBANKA_0802300) and a VPS13 domain-containing protein (PBANKA_1359300) were detected through IP-MS of 1019700::mCherry. However, RNA-binding proteins, such as NAB2 and GBP2, were not detected. In addition, the conserved Plasmodium protein PBANKA_0519900 was detected (see RMgm-5526).

Other mutants


  Tagged: Mutant parasite with a tagged gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_0519900
Gene Model P. falciparum ortholog PF3D7_1036900
Gene productconserved Plasmodium protein, unknown function
Gene product: Alternative name
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 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 parasitehdhfr
Promoter of the selectable markerunknown
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modificationTo generate transgenic parasites expressing GFP-fused PBANKA_0519900 and GFP-fused PBANKA_1359300, the gene-targeting vectors for PBANKA_0519900 and PBANKA_1359300 were prepared by PCR, respectively. The PCR products were annealed to either side of the green fluorescent protein gene (gfp)-mutated human deoxyhypusine synthase (hdhps)-expressing cassette and amplified by PCR using gene-specific primers.
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