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Details of the target gene |
Gene Model of Rodent Parasite |
PBANKA_1212600
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Gene Model P. falciparum ortholog |
PF3D7_1014200
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Gene product | male gamete fusion factor HAP2, putative |
Gene product: Alternative name | HAP2; GCS1, Generative Cell Specific 1 |
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Details of the genetic modification |
Name of the tag | EGFP |
Details of tagging | C-terminal |
Additional remarks: tagging | A 4.2-kb fragment comprising 1.5 kb of upstream sequence and the complete hap2 genomic sequence, except the stop codon, was amplified by PCR from genomic DNA using primers 485 (5′-ATATGGTAC CACGCTACTTATATATAGTGATAACC-3′) and 482 (5′-ATA TGGGCCCTCGCAATGGGGGTATTTTACTTTTAC-3′), inserted into KpnI and ApaI restriction sites of vector p277, upstream of and in frame with an egfp gene, which was followed by 0.5 kb of 3′ untranslated region (UTR) derived from the Pbdhfr/ts gene. The vector was linearized in a unique EcoRV site in the putative hap2 promoter and introduced into a hap2 knockout mutant (RMgm-155). |
Commercial source of tag-antibodies | |
Type of plasmid/construct | Plasmid single cross-over |
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 |
Promoter of the selectable marker | eef1a |
Selection (positive) procedure | WR99210 |
Selection (negative) procedure | No |
Additional remarks genetic modification | A 4.2-kb fragment comprising 1.5 kb of upstream sequence and the complete hap2 genomic sequence, except the stop codon, was amplified by PCR from genomic DNA using primers 485 (5′-ATATGGTAC CACGCTACTTATATATAGTGATAACC-3′) and 482 (5′-ATA TGGGCCCTCGCAATGGGGGTATTTTACTTTTAC-3′), inserted into KpnI and ApaI restriction sites of vector p277, upstream of and in frame with an egfp gene, which was followed by 0.5 kb of 3′ untranslated region (UTR) derived from the Pbdhfr/ts gene. The vector was linearized in a unique EcoRV site in the putative hap2 promoter and introduced into a hap2 knockout mutant (RMgm-155). |
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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