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Details of the target gene |
Gene Model of Rodent Parasite |
PBANKA_1216700
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Gene Model P. falciparum ortholog |
PF3D7_0321800
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Gene product | WD repeat-containing protein, putative |
Gene product: Alternative name | WIG1, WD repeat protein Important for Gametogenesis 1 |
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Details of the genetic modification |
Inducable system used | No |
Additional remarks inducable system |
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Type of plasmid/construct used | (Linear) plasmid double cross-over |
PlasmoGEM (Sanger) construct/vector used | Yes |
Name of PlasmoGEM construct/vector | - |
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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Partial or complete disruption of the gene | Partial |
Additional remarks partial/complete disruption |
We took advantage of the highly efficient PlasmoGEM vectors to delete the last 217 codons of the WIG1 gene (PBANKA_1216700) in P. berghei.
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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 | No |
Additional remarks genetic modification | Triple HA or KO targeting vectors were generated using phage recombineering in Escherichia coli TSA strain with PlasmoGEM vectors (https://plasmogem.umu.se/pbgem/). For final targeting vectors not available in the PlasmoGEM repository, generation of knock-out and tagging constructs were performed using sequential recombineering and gateway steps as previously described. For each gene of interest (goi), the Zeocin-resistance/Phe-sensitivity cassette was introduced using oligonucleotides goi HA-F x goi HA-R and goi KO-F x goi KO-R for 3xHA, AID/HA tagging and KO targeting vectors, respectively. Insertion of the GW cassette following gateway reaction was confirmed using primer pairs GW1 x goi QCR1 and GW2 x goi QCR2. The modified library inserts were then released from the plasmid backbone using NotI. The GD and triple HA targeting vectors were transfected into the 2.34 line. |
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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