RMgmDB - Rodent Malaria genetically modified Parasites

Summary

RMgm-5237
Malaria parasiteP. berghei
Genotype
DisruptedGene model (rodent): PBANKA_1349800; Gene model (P.falciparum): PF3D7_1335900; Gene product: thrombospondin-related anonymous protein | sporozoite surface protein 2 (sporozoite surface protein 2; SSP2; SSP-2; TRAP)
Phenotype Sporozoite;
Last modified: 12 June 2023, 13:59
  *RMgm-5237
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) : 37306042
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/ResearcherBraumann F, Frischknecht F
Name Group/DepartmentIntegrative Parasitology, Center for Infectious Diseases
Name InstituteHeidelberg University Medical School
CityHeidelberg
CountryGermany
Name of the mutant parasite
RMgm numberRMgm-5237
Principal nametrap(-)
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Phenotype
Asexual blood stageNot different from wild type
Gametocyte/GameteNot different from wild type
Fertilization and ookineteNot different from wild type
OocystNot different from wild type
SporozoiteNot analysed in detail:
- Normal oocyst production with sporozoite formation. Sporozoites are released in the hemocoel but are impaired in salivary gland invasion. No (strongly reduced) salivary gland sporozoite numbers.
- The mutant lacks expression of TRAP. The trap gene has been deleted and does not contain a drug-selectable marker (that has been removed using negative selection with 5-FC). This mutant is used to introduce mutated forms of the trap gene.
Liver stageNot tested
Additional remarks phenotype

Mutant/mutation
The mutant lacks expression of TRAP. The trap gene has been deleted and does not contain a drug-selectable marker (that has been removed using negative selection with 5-FC). This mutant is used to introduce mutated forms of the trap gene.

Published in bioRxiv preprint doi: https://doi.org/10.1101/2022.08.24.505106

Protein (function)
TRAP is a type 1 transmembrane protein, containing two adhesive domains in its extracellular portion, an A-domain of von Willebrand factor and a thrombospondin type I repeat (TSR, TSP). TRAP is located in the micronemes of sporozoites. The protein plays a role in the gliding motility of sporozoites and invasion of host cells.

Phenotype
Not analysed in detail: 
- Normal oocyst production with sporozoite formation. Sporozoites are released in the hemocoel but are impaired in salivary gland invasion. No (strongly reduced) salivary gland sporozoite numbers.
- The mutant lacks expression of TRAP. The trap gene has been deleted and does not contain a drug-selectable marker (that has been removed using negative selection with 5-FC). This mutant is used to introduce mutated forms of the trap gene.

Additional information

Other mutants


  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_1349800
Gene Model P. falciparum ortholog PF3D7_1335900
Gene productthrombospondin-related anonymous protein | sporozoite surface protein 2
Gene product: Alternative namesporozoite surface protein 2; SSP2; SSP-2; TRAP
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 usedYes
Name of PlasmoGEM construct/vectorPbGEM-107890
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/yfcu
Promoter of the selectable markereef1a
Selection (positive) procedurepyrimethamine
Selection (negative) procedure5-fluorocytosine (5-FC)
Additional remarks genetic modificationTRAP knockout (trap(-)) parasites were generated with the PlasmoGem vector PbGEM-107890 using standard protocols. Isogenic trap(-) parasites were subsequently negatively selected with 5-fluorocytosine (1.0 mg/mL in the drinking water) to give rise to selection marker-free trap(-) parasites.
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