RMgmDB - Rodent Malaria genetically modified Parasites

Summary

RMgm-5515
Malaria parasiteP. yoelii
Genotype
Genetic modification not successful
DisruptedGene model (rodent): PY17X_1241800; Gene model (P.falciparum): PF3D7_0523800; Gene product: food vacuole resident transporter 1 (FVRT1, PyDMT1 (Divalent metal transporter 1))
PhenotypeNo phenotype has been described
Last modified: 16 June 2024, 21:20
  *RMgm-5515
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 ≥ 5
Reference (PubMed-PMID number) Reference 1 (PMID number) : 38049852
Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. yoelii
Parent strain/lineP. y. yoelii 17X
Name parent line/clone Not applicable
Other information parent line
Attempts to generate the mutant parasite were performed by
Name PI/ResearcherZhong M, Zhou B
Name Group/DepartmentFaculty of Synthetic Biology
Name InstituteShenzhen Institute of Advanced Technology, Chinese Academy of Sciences
CityShenzhen
CountryChina

  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PY17X_1241800
Gene Model P. falciparum ortholog PF3D7_0523800
Gene productfood vacuole resident transporter 1
Gene product: Alternative nameFVRT1, PyDMT1 (Divalent metal transporter 1)
Details of the genetic modification
Inducable system usedNo
Additional remarks inducable system
Type of plasmid/construct usedCRISPR/Cas9 construct: integration through double strand break repair
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
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

To clarify the physiological function of PyDMT1 in Plasmodium, we attempted to knock out PyDMT1 in P. yoelii17X by CRISPR-Cas9. Nine guide RNAs were designed to cover the Pydmt1 gene, and each guide RNA and homologous recombination arms were independently electro-transfected five times. No Pydmt1 knockout parasites were obtained in all these trials.
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