|Successful modification||The parasite was generated by the genetic modification |
|The mutant contains the following genetic modification(s)||
|Reference (PubMed-PMID number)||
Reference 1 (PMID number) : 20132450|
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|Parent parasite used to introduce the genetic modification|
|Rodent Malaria Parasite||P. berghei|
|Parent strain/line||P. berghei ANKA|
|Name parent line/clone
P. berghei ANKA 2.34
|Other information parent line||P. berghei ANKA 2.34 is a cloned, gametocyte producer line of the ANKA strain (PubMed: PMID: 15137943).|
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|The mutant parasite was generated by|
|Name PI/Researcher||K. Slavic; R. Tewari; S. Krishna|
|Name Group/Department||Centre for Infection, Cellular and Molecular Medicine|
|Name Institute||St. George's University of London|
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|Name of the mutant parasite|
|Is the mutant parasite cloned after genetic modification||Yes|
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|Asexual blood stage||GFP-expression in all blood stages. Localization of PbHT-GFP in the plasma membrane of early and late blood stages (see also 'Additional information')|
|Gametocyte/Gamete||GFP expression in female gametes and ookinetes. Colocalization of PbHT-GFP with protein P28 (PB000865.00.0), a surface marker of the ookinete, female gamete and zygote|
|Fertilization and ookinete||GFP expression in female gametes and ookinetes. Colocalization of PbHT-GFP with protein P28 (PB000865.00.0), a surface marker of the ookinete, female gamete and zygote|
|Oocyst||Analysis of mosquito midgut oocysts, 11 days post infection, showed internal foci of GFP-fluorescence of developing sporoblasts and lower intensity membrane fluorescence signal. Twenty-one days post infection, strong fluorescence signal was detectable from sporulating oocysts and oocyst-derived sporozoites. Similarly, sporozoites released from dissected salivary glands 21 days post infection showed expression of PbHT-GFP.|
|Sporozoite||Twenty-one days post infection, strong GFP-fluorescence signal was detectable from sporulating oocysts and oocyst-derived sporozoites. Similarly, sporozoites released from dissected salivary glands 21 days post infection showed expression of PbHT-GFP.|
|Liver stage||Not tested|
|Additional remarks phenotype|
The mutant expresses a C-terminal GFP-tagged version of the hexose transporter (HT)
HT is a facilitative hexose transporter in the Major Facilitator Superfamily of integral membrane proteins that mediates the uptake of glucose and fructose by the parasite. HTis a single copy gene with no close paralogues. In P. falciparum three other proteins, PFI0955w, PFI0785c and PFE1455w have been annotated with putative sugar transport function, although they have diverged considerably from typical sugar transporters (21%, 13% and 7% amino acid sequence identity compared with PfHT, respectively). Unlike PfHT, PFI0955w and PFI0785c are expressed only late in the asexual cycle.
Unsuccessful attempts to disrupt the ht gene in P. berghei and P. falciparum indicate an essential function during asexual blood stage growth (see RMgm-390).
See also RMgm-603 for reports of unsuccessful attempts to disrupt pbht (pbht1) and analyses of a mutant that expresses a HA-tagged form of PbHT (PbHT1).
See also mutant RMgm-604, in which the endogenous hexose transporter 1 gene (pbht1) is replaced with the P. falciparum ortholog, pfht1.
All blood stages, female gametes, ookinetes, oocysts and sporozoites express PbHT-GFP as determined by GFP-fluorescence analysis.
Unsuccessful attempts to disrupt Pbht indicate an essential function during asexual blood stage growth (see RMgm-390). In the same paper, unsuccessful attempts are reported to disrupt the hexose transporter (Pfht) in P. falciparum.
See also RMgm-603 for analyses of a mutant that expresses a HA-tagged form of PbHT (PbHT1).
Some of the observed fluorescence signal in blood stage parasites is internal and may be associated with perinuclear localization as well as the food vacuole. Observed internal PbHT-GFP signal could be associated with developing PbHT in its trafficking pathway or may come from a small proportion of PbHT-GFP protein that has been mislocalized.
Expression of PbHT-GFP in parasites from all analysed developmental stages was confirmed by Western blotting using an anti-GFP monoclonal antibody
RMgm-390: Unsuccessful attempts to disrupt the Pbht gene
RMgm-603: A mutant in which the endogenous hexose transporter 1 gene (pbht1) is replaced with a HA-tagged copy of pbht1.
RMgm-604: a mutant in which the endogenous hexose transporter 1 gene (pbht1) is replaced with the P. falciparum ortholog, pfht1