RMgmDB - Rodent Malaria genetically modified Parasites

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Summary

RMgm-4742
Malaria parasiteP. berghei
Genotype
MutatedGene model (rodent): PBANKA_0605900; Gene model (P.falciparum): PF3D7_1207400; Gene product: conserved Plasmodium protein, unknown function
Details mutation: The PBANKA_0605900 gene of P. berghei replaced with the ortholog of P. falciparum (PF3D7_1207400)
Transgene
 Transgene not Plasmodium: A fusion of GFP (gfp-mu3) and Luciferase Firefly (LucIAV)
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 Sporozoite; Liver stage;
Last modified: 9 March 2020, 17:12
  *RMgm-4742
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 RMgm-4741
Other information parent lineRMgm-4741 is a mutant in which the P. berghei PBANKA_0605900 gene has been replaced with the positive/negative hdhfr/yfcu selectable marker.
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The mutant parasite was generated by
Name PI/ResearcherPul, FJA (Geurten, FJA), Janse CJ
Name Group/DepartmentLeiden Malaria Research Group, Parasitology
Name InstituteLeiden University Medical Center
CityLeiden
CountryThe Netherlands
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Name of the mutant parasite
RMgm numberRMgm-4742
Principal name2484cl1, 2485cl3
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
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Phenotype
Asexual blood stageNot different from wild type
Gametocyte/GameteNot tested
Fertilization and ookineteNot tested
OocystNot different from wild type
Sporozoite(Strongly) reduced numbers of salivary gland sporozoites
Liver stageNo liver infection (and blood infection) after intravenous injection of sporozoites
Additional remarks phenotype

Mutant/mutation
In the mutant the PBANKA_0605900 gene of P. berghei has been replaced with the ortholog of P. falciparum by the method of GIMO transfection. The P. falciparum gene is introduced in mutant RMgm-4741 in which the P. berghei PBANKA_0605900 gene has been replaced with the positive/negative hdhfr/yfcu selectable marker. The P. falciparum ortholog is under control of the regulatory sequences of P. berghei PBANKA_0605900. In addition the mutant expresses the fusion protein GFP-Luciferase under control of the constitutive eef1a promoter

Protein (function)

Phenotype
The phenotype has not been analysed in detail. Normal numbers of oocyst produced. (Strongly) reduced numbers of salivary gland sporozoites. No liver infection (and blood infection) after intravenous injection of sporozoites. These results indicate that the P. falciparum ortholog cannot complement P. berghei PBANKA_0605900 during sporozoite production and liver infection

Additional information
See also RMgm-4741

Other mutants

  Mutated: Mutant parasite with a mutated gene
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Details of the target gene
Gene Model of Rodent Parasite PBANKA_0605900
Gene Model P. falciparum ortholog PF3D7_1207400
Gene productconserved Plasmodium protein, unknown function
Gene product: Alternative name
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Details of the genetic modification
Short description of the mutationThe PBANKA_0605900 gene of P. berghei replaced with the ortholog of P. falciparum (PF3D7_1207400)
Inducable system usedNo
Short description of the conditional mutagenesisNot available
Additional remarks inducable system
Type of plasmid/construct(Linear) PCR construct 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/yfcu
Promoter of the selectable markereef1a
Selection (positive) procedurepyrimethamine
Selection (negative) procedure5-fluorocytosine (5-FC)
Additional remarks genetic modificationIn the mutant the PBANKA_0605900 gene of P. berghei has been replaced with the ortholog of P. falciparum by the method of GIMO transfection. The P. falciparum gene is introduced in mutant RMgm-4741 in which the P. berghei PBANKA_0605900 gene has been replaced with the positive/negative hdhfr/yfcu selectable marker. The P. falciparum ortholog is under control of the regulatory sequences of P. berghei PBANKA_0605900. In addition the mutant expresses the fusion protein GFP-Luciferase under control of the constitutive eef1a promoter
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 nameA fusion of GFP (gfp-mu3) and Luciferase Firefly (LucIAV)
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Details of the genetic modification
Inducable system usedNo
Additional remarks inducable system
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 parasitegfp (FACS)
Promoter of the selectable markereef1a
Selection (positive) procedurepyrimethamine
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: StudioDrie StudioDrie; S. Mededovic and A. van Wigcheren