RMgmDB - Rodent Malaria genetically modified Parasites

Summary

RMgm-1575
Malaria parasiteP. berghei
Genotype
Genetic modification not successful
DisruptedGene model (rodent): PBANKA_0602600; Gene model (P.falciparum): PF3D7_1203600; Gene product: cytochrome c1 heme lyase, putative (CC1HL)
PhenotypeNo phenotype has been described
Last modified: 18 August 2016, 17:43
  *RMgm-1575
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) Reference 1 (PMID number) : 27520480
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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 P. berghei ANKA cl15cy1
Other information parent lineA reference wild type clone from the ANKA strain of P. berghei (PubMed: PMID: 17406255).
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Attempts to generate the mutant parasite were performed by
Name PI/ResearcherPosayapisit N; Kamchonwongpaisana S
Name Group/DepartmentNational Center for Genetic Engineering and Biotechnology (BIOTEC)
Name InstituteNational Science and Technology Development Agency (NSTDA)
CityThanon Phahonyothin, Tambon Khlong Neung, Amphoe Khlong Luang, Pathum Thani
CountryThailand
  Disrupted: Mutant parasite with a disrupted gene
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Details of the target gene
Gene Model of Rodent Parasite PBANKA_0602600
Gene Model P. falciparum ortholog PF3D7_1203600
Gene productcytochrome c1 heme lyase, putative
Gene product: Alternative nameCC1HL
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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/multiplication.

Malaria parasites possess a de novo heme synthetic pathway, which is dispensable during blood stage development in the mammalian host. The assembly of the two most important hemeproteins, cytochromes c and c1, is mediated by cytochrome heme lyase enzymes. Plasmodium spp. possess two cytochrome heme lyases encoded by separate genes. In P. berghei these genes are PBANKA_143950 and PBANKA_0602600. Evidence is presented that these genes can only be disrupted when the parasite is complemented with a gene copy of the P. falciparum ortholog (see also RMgm-1575). Evidence is presented that four genes in the de novo heme synthesis pathway (PBANKA_1459200; PBANKA_0608000; PBANKA_1140700; PBANKA_0105900) can be disrupted without complementation (see RMgm-1576 - RMgm-1580).
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