RMgmDB - Rodent Malaria genetically modified Parasites

Back to search results

Summary

RMgm-1173

Malaria parasite	P. yoelii
Genotype
Tagged	Gene model (rodent): PY17X_1339000; Gene model (P.falciparum): PF3D7_1471100; Gene product: exported protein 2 (EXP2) Name tag: triple-HA
Phenotype	Asexual bloodstage;

Last modified: 30 July 2015, 16:20

*RMgm-1173

Successful modification	The parasite was generated by the genetic modification
The mutant contains the following genetic modification(s)	Gene tagging
Reference (PubMed-PMID number)	Reference 1 (PMID number) : 25662767
MR4 number
top of page
Parent parasite used to introduce the genetic modification
Rodent Malaria Parasite	P. yoelii
Parent strain/line	P. y. yoelii 17X
Name parent line/clone	Not applicable
Other information parent line
top of page
The mutant parasite was generated by
Name PI/Researcher	Meibalan, E; Burns, J.M.
Name Group/Department	Center for Molecular Parasitology
Name Institute	Department of Microbiology and Immunology
City	Drexel University, Philadelphia PA
Country	USA
top of page
Name of the mutant parasite
RMgm number	RMgm-1173
Principal name	PyExp2-3HA
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modification	Yes
top of page
Phenotype
Asexual blood stage	EXP2 was found to be localized to the Parasitophorous Vacuole Membrane (PVM) of blood stgaes. In addition evidence is presented for a location to (spherical) vesicles in the reticulocyte cytoplasm that appeared to be distinct from the PVM.
Gametocyte/Gamete	Not tested
Fertilization and ookinete	Not tested
Oocyst	Not tested
Sporozoite	Not tested
Liver stage	Not tested
Additional remarks phenotype	Mutant/mutation The mutant expresses a C-terminal 3x-HA-tagged version of EXP2. Protein (function) Plasmodium parasites remodel their vertebrate host cells by translocating hundreds of proteins across an encasing membrane into the host cell cytosol via a putative export machinery termed PTEX (Plasmodium Translocon of EXported protein). EXP2 has have been identified as a member of the PTEX complex. Attempts to disrupt the exp2 gene of P. berghei and P. yoeili were unsuccessful, indicating an essential role of EXP2 in blood stages Phenotype EXP2 was found to be localized to the Parasitophorous Vacuole Membrane (PVM) of blood stgaes. In addition evidence is presented for a location to (spherical) vesicles in the reticulocyte cytoplasm that appeared to be distinct from the PVM. Additional information Other mutants

Tagged: Mutant parasite with a tagged gene

top of page

Details of the target gene

Gene Model of Rodent Parasite

PY17X_1339000

Gene Model P. falciparum ortholog

PF3D7_1471100

Gene product

exported protein 2

Gene product: Alternative name

EXP2

top of page

Details of the genetic modification

Name of the tag

triple-HA

Details of tagging

C-terminal

Additional remarks: tagging

Commercial source of tag-antibodies

Type of plasmid/construct

(Linear) plasmid single cross-over

PlasmoGEM (Sanger) construct/vector used

Modified PlasmoGEM construct/vector used

Plasmid/construct map

Plasmid/construct sequence

Restriction sites to linearize plasmid

Selectable marker used to select the mutant parasite

tgdhfr

Promoter of the selectable marker

pbdhfr

Selection (positive) procedure

pyrimethamine

Selection (negative) procedure

Additional remarks genetic modification

To tag the pyexp2 gene, a single cross-over recombination strategy was used. The pSE02-3HA plasmid containing pyrimethamine resistant Tg-dhfr-ts as a selectable marker was used to generate the tagging construct. A 495 bp fragment from the 3’ end of the pyexp2 gene was amplified from P. yoelii 17X genomic DNA and cloned into pSE02-3HA preceding the triple HA sequence resulting in the construct pSE02-3’Exp2-3HA. Plasmid DNA was isolated as above and linearized by digestion with EcoRI prior to transfection.

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

top of page