Summary

RMgm-544
Malaria parasiteP. berghei
Genotype
Genetic modification not successful
DisruptedGene model (rodent): PBANKA_1008200; Gene model (P.falciparum): PF3D7_1436600; Gene product: cGMP-dependent protein kinase (PKG)
PhenotypeNo phenotype has been described
Last modified: 27 May 2012, 17:20
  *RMgm-544
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) : 20951971
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).
Attempts to generate the mutant parasite were performed by
Name PI/ResearcherR. Tewari, O. Billker
Name Group/DepartmentUniversity of Nottingham
Name InstituteUniversity of Nottingham
CityNottingham
CountryUK

  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_1008200
Gene Model P. falciparum ortholog PF3D7_1436600
Gene productcGMP-dependent protein kinase
Gene product: Alternative namePKG
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 nearly complete
Selectable marker used to select the mutant parasitetgdhfr
Promoter of the selectable markerpbdhfr
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modificationThe gene has been targeted for gene deletion using a construct aimed at integration into the genome by double cross-over homologous recombination.

The unsuccessful attempts to disrupt this gene indicates an essential function during blood stage development.

The gene has been targeted for disruption in a 'kinome-wide' study for deletion of genes encoding Plasmodium protein kinases (protein kinase-like proteins).

See the paper for additional information

See also:
RMgm-325: Unsuccessful attempts to disrupt the pkg gene
RMgm-311: A mutants expressing a GFP-tagged (C-terminal) form of PKG
RMgm-357 for a ‘conditional knock-out mutant’ lacking expression of PKG in liver stages. The Flp/FRT site-specific recombination (SSR) system of yeast has been used to silence PKG expression specifically in liver stages

Disruption of the P. falciparum ortholog has been attempted (Solyakov et al., 2011, Nat Commun, 2:565).
The gene is likely essential for asexual proliferation as integration of the KO vector was not achieved. Accesibility of the locus to recombination was verified by C-terminal tagging.
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 1CCCCGGGCCCGGCAATTTTATCTCATGAGGATTTCAC
Additional information primer 1primer sequence target region 1a
Sequence Primer 2GGGGAAGCTTCCCCATAGCGTTCCATTTGTTCC
Additional information primer 2primer sequence target region 1b
Sequence Primer 3CCCCGAATTCGGGTGCACAGTTGATATATGGGC
Additional information primer 3primer sequence target region 2a
Sequence Primer 4GGGGTCTAGACTTCCTCTTCTTGTTCGATCTG
Additional information primer 4primer sequence target region 2b
Sequence Primer 5
Additional information primer 5
Sequence Primer 6
Additional information primer 6