RMgmDB - Rodent Malaria genetically modified Parasites

Summary

RMgm-4880
Malaria parasiteP. berghei
Genotype
MutatedGene model (rodent): PBANKA_1122100; Gene model (P.falciparum): PF3D7_0623200; Gene product: ferredoxin--NADP reductase (FNR)
Details mutation: FNR promoter replaced by eef1a promoter
Transgene
 Transgene not Plasmodium: GFP (gfp-mut3)
Promoter: Gene model: PBANKA_1133300; Gene model (P.falciparum): PF3D7_1357100; Gene product: elongation factor 1-alpha (eef1a)
3'UTR: Gene model: PBANKA_0719300; Gene product: bifunctional dihydrofolate reductase-thymidylate synthase, putative (dhfr/ts)
Replacement locus: Gene model: PBANKA_0306000; Gene product: 6-cysteine protein (230p)
Phenotype Asexual bloodstage;
Last modified: 30 September 2020, 16:21
  *RMgm-4880
Successful modificationThe parasite was generated by the genetic modification
The mutant contains the following genetic modification(s) Gene mutation, Introduction of a transgene
Reference (PubMed-PMID number) Not published (yet)
MR4 number
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Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. berghei
Parent strain/lineP. berghei ANKA
Name parent line/clone PQ(r)
Other information parent lineThe P. berghei ANKA reference line 507cl1 (RMgmDB-7) which is selected for piperaquine resistance and expresses GFP from the constitutive eef1a promoter (described in PMID 19318094)
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The mutant parasite was generated by
Name PI/ResearcherBara FD, Kiboi D
Name Group/DepartmentDepartment of Biochemistry
Name InstituteJomo Kenyatta University of Agriculture and Technology (JKUAT)
CityNairobi
CountryKenya
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Name of the mutant parasite
RMgm numberRMgm-4880
Principal namePQ(R)_FNR_OE
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
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Phenotype
Asexual blood stageReduced asexual growth/multiplication in mice (47%)
Gametocyte/GameteNot tested
Fertilization and ookineteNot tested
OocystNot tested
SporozoiteNot tested
Liver stageNot tested
Additional remarks phenotype

Mutant/mutation
The mutant expresses a mutated FNR gene locus where the FNR promoter has been replaced by the eef1a promoter (PBANKA_1133300), using a PlasmoGem 'over-expression' construct
(bioRxiv preprint doi: https://doi.org/10.1101/833145. this version posted September 14, 2020)

Protein (function)

Phenotype
Reduced asexual growth/multiplication in mice (47%)

Additional information
To confirm the overexpression of the FNR, we quantified the mRNA expression in PQR_FNR_OE line relative to the PQR parasites. The results indicated a 5-fold (p<0.0046) increase in the mRNA amount of the FNR gene in the PQR_FNR_OE parasites

Over-expression of FNR in the PQ(R) parasites abolished the impact of the chemosensitizers probenecid, verapamil, or cyproheptadine on lumefantrine (LM) activity, and restored the susceptibility of the PQ(R) parasites to LM.

Other mutants

  Mutated: Mutant parasite with a mutated gene
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Details of the target gene
Gene Model of Rodent Parasite PBANKA_1122100
Gene Model P. falciparum ortholog PF3D7_0623200
Gene productferredoxin--NADP reductase
Gene product: Alternative nameFNR
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Details of the genetic modification
Short description of the mutationFNR promoter replaced by eef1a promoter
Inducable system usedNo
Short description of the conditional mutagenesisNot available
Additional remarks inducable system
Type of plasmid/construct(Linear) plasmid double cross-over
PlasmoGEM (Sanger) construct/vector usedYes
Name of PlasmoGEM construct/vectorPbGEM-456502
Modified PlasmoGEM construct/vector usedNo
Plasmid/construct map
Plasmid/construct sequence
Restriction sites to linearize plasmid
Selectable marker used to select the mutant parasitehdhfr/yfcu
Promoter of the selectable markereef1a
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 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
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  Transgene: Mutant parasite expressing a transgene
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Type and details of transgene
Is the transgene Plasmodium derived Transgene: not Plasmodium
Transgene nameGFP (gfp-mut3)
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Details of the genetic modification
Inducable system usedNo
Additional remarks inducable system
Type of plasmid/construct(Linear) plasmid single 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 parasitegfp (FACS)
Promoter of the selectable markereef1a
Selection (positive) procedureFACS (flowsorting)
Selection (negative) procedureNo
Additional remarks genetic modification
Additional remarks selection procedure
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Other details transgene
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Promoter
Gene Model of Parasite PBANKA_1133300
Gene Model P. falciparum ortholog PF3D7_1357100
Gene productelongation factor 1-alpha
Gene product: Alternative nameeef1a
Primer information details of the primers used for amplification of the promoter sequence  Click to view information
Primer information details of the primers used for amplification of the promoter sequence  Click to hide information
Sequence Primer 1
Additional information primer 1
Sequence Primer 2
Additional information primer 2
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3'-UTR
Gene Model of Parasite PBANKA_0719300
Gene productbifunctional dihydrofolate reductase-thymidylate synthase, putative
Gene product: Alternative namedhfr/ts
Primer information details of the primers used for amplification the 3'-UTR sequences  Click to view information
Primer information details of the primers used for amplification the 3'-UTR sequences  Click to hide information
Sequence Primer 1
Additional information primer 1
Sequence Primer 2
Additional information primer 2
Insertion/Replacement locus
Replacement / InsertionReplacement locus
Gene Model of Parasite PBANKA_0306000
Gene product6-cysteine protein
Gene product: Alternative name230p
Primer information details of the primers used for amplification of the target sequences  Click to view information
Primer information details of the 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
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Conceptual design of the database: Dr. C.J. Janse (Leiden Malaria Research Group, Leiden, The Netherlands).
Technical design and realization: S. Mededovic and A. van Wigcheren