RMgmDB - Rodent Malaria genetically modified Parasites


Malaria parasiteP. berghei
TaggedGene model (rodent): PBANKA_1413600; Gene model (P.falciparum): PF3D7_1315100; Gene product: serine/threonine protein kinase PK9
Name tag: triple-HA
Phenotype Asexual bloodstage; Sporozoite; Liver stage;
Last modified: 26 May 2022, 17:51
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) : 35490588
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/ResearcherNarwal SK, Mishra S
Name Group/DepartmentDivision of Molecular Microbiology and Immunology
Name InstituteCSIR-Central Drug Research Institute
Name of the mutant parasite
RMgm numberRMgm-5207
Principal namePk9-3xHA
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Asexual blood stageCytoplasmic expression of in Pk9-3xHA in blood stages
Gametocyte/GameteNot tested
Fertilization and ookineteNot tested
OocystNot tested
SporozoiteCytoplasmic expression of Pk9-3xHA in sporozoites
Liver stageCytoplasmic expression of Pk9-3xHA in (late) liver stages
Additional remarks phenotype

The mutant expresses a C-terminal 3xHA-tagged version of PK9

Protein (function)
Previous studies have shown that functionally Pk9 can be designated as a serine-threonine kinase (STK) owing to its ability to phosphorylate E2 conjugation enzyme at serine 106 residue of Ubc13, although it does not cluster with any established eukaryotic protein kinase groups and hence is referred to as orphan kinase..

Cytoplasmic expression of Pk9-3xHA in blood stages, sporozoite stages and (late) liver stages

See also mutant RMgm-5206 lacking expression of PK9.

Additional information
In this study also evidence is presented that that phosphorylation of the regulatory serine 106 of Ubc13 is essential for the completion of the parasite’s erythrocytic cycle. However,  phosphorylated form of Ubc13 was observed in Pk9 knockout parasites (RMgm-5206).

The process of ubiquitylation typically requires three enzymes: E1 (ubiquitin-activating enzyme, Uba), E2 (ubiquitin-conjugating enzyme, Ubc), and E3 (ubiquitin-protein ligase). Amongst Ubcs, Ubc13 specifically catalyzes the formation of K63-linked ubiquitin chains that are critical to signaling in inflammatory and DNA damage response pathways.

See also mutant RMgm-5208 with a mutated UNC13 (the phosphorylating serine 106 replaced with with non-phosphorylatable alanine).

Other mutants

  Tagged: Mutant parasite with a tagged gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_1413600
Gene Model P. falciparum ortholog PF3D7_1315100
Gene productserine/threonine protein kinase PK9
Gene product: Alternative name
Details of the genetic modification
Name of the tagtriple-HA
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 markereef1a
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modificationFor the construction of a Pk9-3XHA tagging plasmid, two fragments F1 and F2 corresponding to 0.76 kb Pk9 ORF and 0.6 kb Pk9 3′UTR were amplified using primer pairs 1020/1021 and 1018/1019 respectively. Fragments F1 and F2 were cloned sequentially into pBC-3XHA-hDHFR plasmid at XhoI/BglII and NotI/AscI restriction sites respectively
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