RMgmDB - Rodent Malaria genetically modified Parasites

Summary

RMgm-5125
Malaria parasiteP. berghei
Genotype
Genetic modification not successful
DisruptedGene model (rodent): PBANKA_1033200; Gene model (P.falciparum): PF3D7_1409300; Gene product: DNA damage-inducible protein 1, putative
PhenotypeNo phenotype has been described
Last modified: 13 December 2021, 13:41
  *RMgm-5125
Successful modificationThe gene/parasite could not be changed/generated by the genetic modification.
The following genetic modifications were attempted Gene disruption
Number of attempts to introduce the genetic modification Unknown
Reference (PubMed-PMID number) Not published (yet)
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
Attempts to generate the mutant parasite were performed by
Name PI/ResearcherTanneru N, Sijwali PS
Name Group/DepartmentCSIR-Centre for Cellular and Molecular Biology
Name InstituteCSIR-Centre for Cellular and Molecular Biology
CityHyderabad
CountryIndia

  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_1033200
Gene Model P. falciparum ortholog PF3D7_1409300
Gene productDNA damage-inducible protein 1, putative
Gene product: Alternative name
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 usedNo
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
Promoter of the selectable markerunknown
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modificationPublished in: bioRxiv preprint doi: https://doi.org/10.1101/2021.10.29.466443

The unsuccessful attempts to disrupt this gene indicate an essential function during asexual blood stage growth/multiplication.

The PbDDI1 gene was targeted for knock-out and knock-down using double cross-over homologous recombination approach.
For the knock-out constructs, the 5'-UTR (flank 1) and 3’-UTR (flank 2) of PbDDI1 were amplified from P. berghei genomic DNA using PbDdi1- Fl1F/ PbDdi1-Fl1R and PbDdi1-Fl2F/PbDdi1-Fl2R primer sets, respectively. The flank 1 and flank 2 were cloned into the HB-DJ1KO plasmid at NotI-KpnI and AvrII-KasI sites, respectively, to obtain HB-PbDDI-(FL1+FL2) plasmid. The GFP coding sequence was excised from pGT-GFPbsc with KpnI-XhoI and subcloned into the similarly digested HB-PbDDI- (FL1+FL2) to obtain HB-pbDDIKO plasmid.
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