Summary

RMgm-160
Malaria parasiteP. berghei
Genotype
Genetic modification not successful
DisruptedGene model (rodent): PBANKA_0911700; Gene model (P.falciparum): PF3D7_1136900; Gene product: subtilisin-like protease 2 (SUB2)
PhenotypeNo phenotype has been described
Last modified: 24 February 2009, 16:02
  *RMgm-160
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 3
Reference (PubMed-PMID number) Reference 1 (PMID number) : 14678331
Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. berghei
Parent strain/lineP. berghei ANKA
Name parent line/clone P. berghei ANKA cl15cy1
Other information parent line
Attempts to generate the mutant parasite were performed by
Name PI/ResearcherP. Uzureau; C.J. Janse; A.P. Waters; C. Braun Breton
Name Group/DepartmentUnité de Biologie des Interactions Hôte-Parasite
Name InstituteInstitut Pasteur
CityParis
CountryFrance

  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_0911700
Gene Model P. falciparum ortholog PF3D7_1136900
Gene productsubtilisin-like protease 2
Gene product: Alternative nameSUB2
Details of the genetic modification
Inducable system usedNo
Additional remarks inducable system
Type of plasmid/construct usedPlasmid 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 genePartial
Additional remarks partial/complete disruption The construct was specifically designed to lead to the loss of 400 bp coding for part of the subtilisin catalytic domain including the active serine residue (see Supplementary material, Fig S2 in: Uzureau et al.,2004,Cell Microbiol. 6:65-78).
Selectable marker used to select the mutant parasitetgdhfr
Promoter of the selectable markerpbdhfr
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modificationSUB2 is a subtilisin-like serine protease. In Plasmodium three subtilisin-like proteases (SUB1-3) have been identified. SUB2 has been shown to be associated with dense granules and/or micronemes, the contents of which are released during erythrocyte invasion. It has been proposed to function as a 'sheddase', involved in the release of MSP1 and AMA1 during invasion (Harris et al., 2005, PloS Pathogens, 3, e29).
The failure to disrupt the gene indicates an essential function in the blood stages.
The generation of a mutant containing the endogenous sub2 gene tagged with c-myc (RMgm-161) demonstrates that the sub2 locus is accessible to genetic modification (Uzureau et al.,2004,Cell Microbiol. 6:65-78).
See also GenBank accession numbers AJ242629 and AF145052 for the correct sequence of the gene encoding SUB2.
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