RMgmDB - Rodent Malaria genetically modified Parasites

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Summary

RMgm-4425
Malaria parasiteP. yoelii
Genotype
Genetic modification not successful
DisruptedGene model (rodent): PY17X_1121600; Gene model (P.falciparum): PF3D7_0621300; Gene product: mRNA-binding protein PUF3 (PUF3)
PhenotypeNo phenotype has been described
Last modified: 5 March 2018, 17:00
  *RMgm-4425
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) : 29487181
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Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. yoelii
Parent strain/lineP. y. yoelii 17XNL
Name parent line/clone Not applicable
Other information parent line
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Attempts to generate the mutant parasite were performed by
Name PI/ResearcherLiang X, Lindner SE, Cui L
Name Group/DepartmentDepartment of Entomology
Name InstitutePennsylvania State University
CityUniversity Park
CountryUSA
  Disrupted: Mutant parasite with a disrupted gene
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Details of the target gene
Gene Model of Rodent Parasite PY17X_1121600
Gene Model P. falciparum ortholog PF3D7_0621300
Gene productmRNA-binding protein PUF3
Gene product: Alternative namePUF3
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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
Promoter of the selectable markerunknown
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 the paper evidence is presented that:
- Secondary structure prediction suggested that the RNA-binding domains of Puf3s consisted of 11 pumilio repeats, similar to those in human Puf-A and yeast Puf6, which are involved in ribosome biogenesis.
- Neither pfpuf3 nor pypuf3 could be genetically disrupted, suggesting they may be essential for the intraerythrocytic developmental cycle
- Nuclear, nucleolar location
- PyPuf3 changed its localization in mosquito and liver stages from nucleolar to cytosolic puncta.
- Affinity purification of PTP-tagged variant of PfPuf3 revealed an association with 31 proteins associated with the 60S ribosome, and an enrichment of 28S rRNA and internal transcribed spacer 2 sequences. Taken together, these results suggest an essential function of PfPuf3 in ribosomal biogenesis
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