RMgmDB - Rodent Malaria genetically modified Parasites

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Summary

RMgm-5402
Malaria parasiteP. berghei
Genotype
Genetic modification not successful
DisruptedGene model (rodent): PBANKA_0922200; Gene model (P.falciparum): PF3D7_1126100; Gene product: autophagy-related protein 7, putative (Atg7)
PhenotypeNo phenotype has been described
Last modified: 5 March 2024, 22:27
  *RMgm-5402
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) Not published (yet)
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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 Not applicable
Other information parent linePlasmodium berghei ANKA (MRA 311) was obtained from BEI resources, USA
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Attempts to generate the mutant parasite were performed by
Name PI/ResearcherMishra A, Mishra S
Name Group/DepartmentDivision of Molecular Microbiology and Immunology
Name InstituteCSIR-Central Drug Research Institute
CityLucknow
CountryIndia
  Disrupted: Mutant parasite with a disrupted gene
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Details of the target gene
Gene Model of Rodent Parasite PBANKA_0922200
Gene Model P. falciparum ortholog PF3D7_1126100
Gene productautophagy-related protein 7, putative
Gene product: Alternative nameAtg7
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Details of the genetic modification
Inducable system usedNo
Additional remarks inducable system
Type of plasmid/construct used(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
Partial or complete disruption of the geneComplete
Additional remarks partial/complete disruption
Selectable marker used to select the mutant parasitehdhfr/yfcu
Promoter of the selectable markereef1a
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modificationThe unsuccessful attempts to disrupt this gene indicate an essential function during asexual blood stage growth.

In an attempt to generate direct knockout of PbAtg7, two fragments, F3 (0.54 kb) and F4 (0.61 kb), were amplified using primers 1321/1322 and 1323/1324 and sequentially cloned and inserted into pBC-GFP-yFCU-hDHFR at blunt-ended SalI and NotI/AscI sites, respectively. The plasmid was linearized with XhoI/AscI restriction digestion and transfected into P. berghei ANKA schizonts
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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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