Summary

RMgm-64
Malaria parasiteP. berghei
Genotype
Genetic modification not successful
DisruptedGene model (rodent): PBANKA_0524200; Gene model (P.falciparum): Not available; Gene product: early transcribed membrane protein; small exported protein (SEP2; ETRAMP4; Pbsep2)
PhenotypeNo phenotype has been described
Last modified: 27 July 2015, 18:22
  *RMgm-64
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 2
Reference (PubMed-PMID number) Not published (yet)
Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. berghei
Parent strain/lineP. berghei ANKA
Name parent line/clone 8417HP
Other information parent lineA reference wild type clone from the ANKA strain of P. berghei (Janse et al., Exp. Parasitol. 68, 274-282).
Attempts to generate the mutant parasite were performed by
Name PI/ResearcherPace, T; Ponzi, M
Name Group/DepartmentLaboratorio di Biologia Cellulare
Name InstituteIstituto Superiore di Sanita
CityRome
CountryItaly

  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_0524200
Gene Model P. falciparum ortholog Not available
Gene productearly transcribed membrane protein; small exported protein
Gene product: Alternative nameSEP2; ETRAMP4; Pbsep2
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 parasitepbdhfr
Promoter of the selectable markerpbdhfr
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modificationIn P. berghei, this gene belongs to a family of three related genes (encoding so-called SEP proteins; sep1-3), located in the subtelomeric regions of chromosomes 5 (Birago et al., Mol. Biochem. Parasitol. (2003) 126, 209-18).
The SEP proteins show homology to members of the ETRAMP family of proteins of P. falciparum but the orthologous relationship of the different members is unclear (ortholog of etramp4, PFD1120c?).

The gene encodes encodes an integral membrane protein. The protein contains a predicted signal peptide at the NH(2)-terminus, an internal hydrophobic region and a polymorphic, low-complexity region at the carboxy-terminus (Birago et al., Mol. Biochem. Parasitol. (2003) 126, 209-18).

The failure to disrupt sep2 (sep-bis) has not been published yet. Two transfections has been performed using a BioRad electroporation device(pers. comm. M. Ponzi, T. Pace).

The coding region of this gene as confirmed by ESTs is as follows (pers. comm. Dr. T. Pace):
MKLAKALYFVAFLLAIKVLTPGSNNYVEAKPANSKKVTKSGDNAFIKKIKNNKAAFISTLAA
TVALAIATTFGVMHYQNNGGFEKSPWANKIDKNKSLPKKKTDNSPPSKGNLGNTSQKRNPISF
TPISDGYKPSSSPNNNSKRTPTAPYTVKLN

The two other sep genes, sep1 (PB100384.00.0; PF11_0039; early transcribed membrane protein, etramp) and sep3 (PB401867.00.0; sep-ter) have been targeted for gene disruption.
See RMgm-16 and RMgm-18 for mutants containing targeted disruptions of sep1
See RMgm-65 for more information on sep3.
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 1gaccgaattccatgtatatacccaagacacataaac
Additional information primer 1KObis-sin-for
Sequence Primer 2gaccctcgagtgcataatatcagcatagctaaaac
Additional information primer 2KObis-sin-rev
Sequence Primer 3gaccggtaccccaaaccaataagacgacaatg
Additional information primer 3KObis-dx-for
Sequence Primer 4gaccgcggccgcgcataattgtagtactattattgcg
Additional information primer 4KObis-dx-rev
Sequence Primer 5
Additional information primer 5
Sequence Primer 6
Additional information primer 6