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Details of the target gene |
Gene Model of Rodent Parasite |
PBANKA_0417600
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Gene Model P. falciparum ortholog |
PF3D7_0903800
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Gene product | LCCL domain-containing protein |
Gene product: Alternative name | CCp4; LAP6 |
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Details of the genetic modification |
Name of the tag | EGFP |
Details of tagging | C-terminal |
Additional remarks: tagging | |
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 | pbdhfr |
Selection (positive) procedure | pyrimethamine |
Selection (negative) procedure | No |
Additional remarks genetic modification | To achieve GFP-tagging of PbLAP6 we adopted a strategy of single crossover homologous recombination. A ca. 1.9 kb fragment of pblap6 corresponding to the 3-part of the coding sequence was PCR amplified from genomic DNA with primers P1 (ACGAAGTTATCAGTCGACAGCCCCAGTTCAGACATAAAC) and P2 (ATGAGGGCCCCTAAGCTTTCTTTATGAGGAATAAATAAAATGTTTTTAAAC) and introduced into SalI/HindIII-digested pDNR-EGFP via in-fusion cloning(Takara Biotech) to give plasmid pDNR-PbLAP6/EGFP. The pblap6/egfp specific sequence was then transferred to pLP-hDHFR via cre-loxp recombination to give plasmid pLP-PbLAP6/EGFP |
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 | ACGAAGTTATCAGTCGACAGCCCCAGTTCAGACATAAAC |
Additional information primer 1 | P1 |
Sequence Primer 2 | ATGAGGGCCCCTAAGCTTTCTTTATGAGGAATAAATAAAATGTTTTTAAAC |
Additional information primer 2 | P2 |
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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