Mutant/mutation
The mutant lacks expression of CDPK3 (calcium-dependent protein kinase 3) and expresses GFP under the constitutive HSP70 promoter.
This mutant has been generated by a genetic cross between mutant RMgm-165 lacking expression of CDPK3 and mutant RMgm-166 that expresses GFP (see 'Additional information: genetic modification).

Protein (function)
CDPK3 belongs to an expanded family of Ca²⁺ dependent protein kinases (CDPKs). CDPKs combine an amino-terminal serine/threonine kinase domain and a carboxy-terminal calmodulin-like domain, composed of four EF hands, in the same molecule. In plants, CDPKs translate Ca²⁺ signals generated by external stimuli into cellular responses, thereby regulating cell division and differentiation, the development of tolerance to stress stimuli and the specific defense responses to pathogens.

Phenotype
The phenotype analyses indicate a role of CDPK3 in the transformation of the mature ookinete into the oocyst. Specifically, ookinetes fail to bind and traverse the cell of the midgut wall.
This mutant has been generated by a genetic cross between mutant RMgm-165 lacking expression of CDPK3 and mutant RMgm-166 that expresses GFP (see 'Additional information: genetic modification). See also RMgm-165 for a description of the phenotype of the mutant lacking expression of CDPK3. Electron microscopic analyses of mutant RMgm-165 demonstrated that the disruptant ookinetes could not access midgut epithelial cells by traversing the layer covering the cell surface.
The GFP-fluorescent ookinetes have been used to analyse the in vitro motility behavior of ookinetes. Ookinetes are severely impaired in the ability to migrate in vitro through a gel structure (Matrigel; see 'Additional information'). Ookinetes were able to move on the gel surface.

Additional information
The GFP-fluorescent ookinetes have been used to analyse the in vitro motility behavior of ookinetes. Cultured GFP-expressing wild-type or mutant ookinetes were inoculated to the top of a layer of gelled basement membrane matrix (Matrigel) in a 24 well plate and were allowed to migrate. After 8 h the number of ookinetes that entered the Matrigel and the depth they reached were measured by optical and confocal microscopy. Soon after inoculation, almost all wild-type ookinetes entered the gel and started moving towards the bottom of the layer. In contrast most mutant ookinetes remained on the gel surface. The small proportion of disruptant ookinetes (6%) that succeeded in entering the gel remained near the surface. These results indicated that mutant ookinetes are impaired in the ability to migrate through a gel structure, which might explain the failure to cross the layer covering the surface of the midgut epithelium.
Real time images of mutant ookinetes showed that they are able to move on the gel surface. This indicates that without CDPK3 ookinetes have a surface motility that is distinguishable from their ability to traverse a gel.

An independent mutant lacking expression of CDPK3, RMgm-154, has been generated which show a comparable defect in ookinete to oocyst transition. However, instead of the specific defect in traversal of the layer covering the epithelial cells, a more general defect in motility of the ookinetes was observed, suggesting a role for CDPK3 in regulating productive gliding motility of ookinetes.

Disruption of the P. falciparum ortholog has been attempted (Solyakov et al., 2011, Nat Commun, 2:565).
The gene is likely essential for asexual proliferation. After transfection with a KO vector a weak PCR signal diagnostic for integration was observed, indicating that integration does transiently occur but parasites with a disrupted locus do not persist. Cloning will be required to validate this interpretation for this gene.

Other mutants
RMgm-154: An independent mutant lacking expression of CDPK3
RMgm-165: A mutant lacking expression of CDPK3 which has been used to genereate muatnt RMgm-168 by genetic crossing with mutant RMgm-166,  that expresses GFP
RMgm-12: A mutant lacking expression CDPK4