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Details of the target gene |
Gene Model of Rodent Parasite |
PY17X_0515900
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Gene Model P. falciparum ortholog |
PF3D7_1030900
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Gene product | ookinete surface protein P28 |
Gene product: Alternative name | P28, Pfs28 |
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Details of the genetic modification |
Name of the tag | mCherry |
Details of tagging | C-terminal |
Additional remarks: tagging | |
Commercial source of tag-antibodies | |
Type of plasmid/construct | CRISPR/Cas9 construct: integration through double strand break repair |
PlasmoGEM (Sanger) construct/vector used | No |
Modified PlasmoGEM construct/vector used | No
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Plasmid/construct map |
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Plasmid/construct sequence |
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Restriction sites to linearize plasmid |
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Selectable marker used to select the mutant parasite | hdhfr/yfcu |
Promoter of the selectable marker | eef1a |
Selection (positive) procedure | pyrimethamine |
Selection (negative) procedure | 5-fluorocytosine (5-FC) |
Additional remarks genetic modification | For tagging P28 the CRISPR/Cas9 method was used as described for mutant RMgm-1095 with a slight modification of the pYCm vector. In this vector the original hdhfr slectable marker was replaced by the hdhfr-yfcu selectable marker. This allows for killing mutated parasites that still contain remaining episomal plasmids by 5-FC treatment of mice. Removal of the episomal plasmid is essential for subsequent mutations in the same parasite using the CRISPR/Cas9 method. |
Additional remarks selection procedure | |
Primer information: Primers used for amplification of the target sequences
Primer information: Primers used for amplification of the target sequences
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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