Summary

RMgm-918
Malaria parasiteP. berghei
Genotype
DisruptedGene model (rodent): PBANKA_1358000; Gene model (P.falciparum): PF3D7_1345100; Gene product: thioredoxin 2 (TRX2)
Phenotype Asexual bloodstage;
Last modified: 30 July 2013, 21:52
  *RMgm-918
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) : 23869529
MR4 number
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 lineA reference wild type clone from the ANKA strain of P. berghei (PubMed: PMID: 17406255).
The mutant parasite was generated by
Name PI/ResearcherMatthews, K; de Koning-Ward, T.F.
Name Group/DepartmentSchool of Medicine
Name InstituteDeakin University
CityWaurn Ponds, Victoria
CountryAustralia
Name of the mutant parasite
RMgm numberRMgm-918
Principal nameΔPbTRX2
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Phenotype
Asexual blood stageBlood stages show a delayed growth in mice. Evidence is presented for a prolonged cell cycle of asexual blood stages, mainly affecting the development of ring forms and schizonts. The number of merozoites per schizont was comparable to that of wild type schizonts.
Gametocyte/GameteNot tested
Fertilization and ookineteNot tested
OocystNot tested
SporozoiteNot tested
Liver stageNot tested
Additional remarks phenotype

Mutant/mutation
The mutant lacks expression of TRX2.

Protein (function)
Thioredoxins act as both reducing agents and protein disulfide reductases.
P. falciparum contains three thioredoxins - Trx1, Trx2, Trx3 and two thioredoxin-like proteins (Tlp1 and Tlp2). Trx1 and Tlp1 are cytosolic, Trx3 is localized to the endoplasmic reticulum, while Tlp2 is mitochondrial. Thioredoxin 2 is a 19 kDa protein with an ER signal sequence and is part of the so-called translocon complex (PTEX) which is a parasite derived multi-protein machinery resident in parasite parasitophorous vacuolar membrane (PVM), responsible for protein secretion into host cells
Plasmodium parasites remodel their vertebrate host cells by translocating hundreds of proteins across an encasing membrane into the host cell cytosol via a putative export machinery termed PTEX (Plasmodium Translocon of EXported protein). HSP101 (PbANKA_094120), PTEX150 (PbANKA_100850), EXP2 (PbANKA_133430), PTEX88 (PbANKA_094130) and TRX2 (PbANKA_135800) have been identified as members of the PTEX complex.
These proteins are also expressed  in early gametocytes, mosquito and liver stages. Although amenable to genetic tagging, HSP101, PTEX150, EXP2 and PTEX88 could not be genetically deleted in P. berghei, in keeping with the obligatory role this complex is postulated to have in maintaining normal blood-stage growth. In contrast, the putative thioredoxin-like protein TRX2 could be deleted, with knockout parasites displaying reduced grow-rates, both in vivo and in vitro.

Phenotype
Blood stages show a delayed growth in mice. Evidence is presented for a prolonged cell cycle of asexual blood stages, mainly affecting the development of ring forms and schizonts. The number of merozoites per schizont was comparable to that of wild type schizonts.

Additional information
Infections with the mutant parasite do not induce experimental cerebral malaria (ECM) in C57Bl6 mice.

Other mutants


  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_1358000
Gene Model P. falciparum ortholog PF3D7_1345100
Gene productthioredoxin 2
Gene product: Alternative nameTRX2
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 geneComplete
Additional remarks partial/complete disruption
Selectable marker used to select the mutant parasitetgdhfr
Promoter of the selectable markerpbdhfr
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modification
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 1tagggcccTTCCTAGTAAAATTACGACCTTATC
Additional information primer 1S5F: PbTRX2 5’ UTR (ApaI)
Sequence Primer 2gttccgcggCGCTAATATGTGCTTCATGATGA
Additional information primer 2S6R: PbTRX2 5’ UTR (SacII)
Sequence Primer 3cgcgaattcGAATAAGCTATGATATTAGGT
Additional information primer 3T16F: PbTRX2 3’ UTR (EcoRI)
Sequence Primer 4cgcactagtCAGAGGAGTGTCAACTGGTAT
Additional information primer 4T17R: PbTRX2 3’ UTR (SpeI)
Sequence Primer 5
Additional information primer 5
Sequence Primer 6
Additional information primer 6