RMgmDB - Rodent Malaria genetically modified Parasites

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Summary

RMgm-4123
Malaria parasiteP. chabaudi
Genotype
Transgene
Transgene not Plasmodium: Firefly luciferase
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)
Insertion locus: Gene model: PCHAS_0308200; Gene product: 6-cysteine protein (P230p)
Phenotype Asexual bloodstage;
Last modified: 10 February 2017, 10:54
  *RMgm-4123
Successful modificationThe parasite was generated by the genetic modification
The mutant contains the following genetic modification(s) Introduction of a transgene
Reference (PubMed-PMID number) Reference 1 (PMID number) : 28155887
MR4 number
Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. chabaudi
Parent strain/lineP. c.chabaudi AS
Name parent line/clone Not applicable
Other information parent lineCloned lines of Plasmodium chabaudi chabaudi AS were originally obtained from David Walliker, University of Edinburgh, UK and subsequently passaged through mice by injection of infected red blood cells (iRBC) at the MRC National Institute for Medical Research, UK
The mutant parasite was generated by
Name PI/ResearcherLin J, Langhorne J
Name Group/DepartmentMalaria Immunology laboratory
Name InstituteFrancis Crick Institute
CityLondon
CountryUK
Name of the mutant parasite
RMgm numberRMgm-4123
Principal namePccASluc230p
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Phenotype
Asexual blood stageLuciferase expression in blood stages as shown by in vivo imaging of luminescence signals in mice and isolated organs infected with the transgenic parasites.
Gametocyte/GameteNot tested
Fertilization and ookineteNot tested
OocystNot tested
SporozoiteNot tested
Liver stageNot tested
Additional remarks phenotype

Mutant/mutation
The mutant contains the luciferase gene under control of the P. berghei elongation factor 1 alpha (eef1a) promoter integrated into the 230p gene locus

Protein (function)

Phenotype
Luciferase expression in blood stages as shown by in vivo imaging of luminescence signals in mice and isolated organs infected with the transgenic parasites.

Additional information
Parasite loads are determined by in vivo imaging of bioluminescence in whole mice and isolated organs of mice infected with P. c. chabaudi AS and P. c. chabaudi CB. Sequestration of parasites is observed in spleens, liver and lungs. Higher parasite loads in lungs of mice infected with the more virulent line P. c. chabaudi CB compared to P. c. chabaudi AS.

Other mutants
RMgm-4124 - P. c. chabaudi CB containing  the luciferase gene under control of the P. berghei elongation factor 1 alpha (eef1a) promoter integrated into the 230p gene locus
RMgm-937 - P. c. chabaudi AS containing  the luciferase gene under control of the P. berghei elongation factor 1 alpha (eef1a) promoter integrated into the small subunit (ssu) ribosomal RNA locus


  Transgene: Mutant parasite expressing a transgene
Type and details of transgene
Is the transgene Plasmodium derived Transgene: not Plasmodium
Transgene nameFirefly luciferase
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 parasitetgdhfr
Promoter of the selectable markerpbdhfr
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modificationP. c. chabaudi AS expressing luciferase under the control of the constitutive promoter eef1a was generated by transfection with the construct pPc-LUC230p, targeting the neutral P230p locus (PCHAS_0308200). The construct pPc-LUC230p was modified from pPc-LUCCAM8 (RMgm-937) by replacing the P. chabaudi SSU targeting region with 230p targeting region, Chab03 277001-278950, generated by gene synthesis (Genewiz LLC, NJ, USA).
Additional remarks selection procedure
Other details transgene
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
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 / InsertionInsertion locus
Gene Model of Parasite PCHAS_0308200
Gene product6-cysteine protein
Gene product: Alternative nameP230p
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