SummaryRMgm-4397
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Successful modification | The parasite was generated by the genetic modification |
The mutant contains the following genetic modification(s) | Gene tagging |
Reference (PubMed-PMID number) |
Reference 1 (PMID number) : 29233900 |
MR4 number | |
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Parent parasite used to introduce the genetic modification | |
Rodent Malaria Parasite | P. yoelii |
Parent strain/line | P. y. yoelii 17XNL |
Name parent line/clone | Not applicable |
Other information parent line | |
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The mutant parasite was generated by | |
Name PI/Researcher | Zhang C; Yuan J |
Name Group/Department | State Key Laboratory of Cellular Stress Biology, Innovation Center for Cell Signaling Network, Schoo |
Name Institute | Xiamen University |
City | Xiamen, Fujian |
Country | China |
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Name of the mutant parasite | |
RMgm number | RMgm-4397 |
Principal name | Pyap2-g3 HA-tagged |
Alternative name | |
Standardized name | |
Is the mutant parasite cloned after genetic modification | Yes |
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Phenotype | |
Asexual blood stage | We tagged PyAP2-G3 with 6xHA at the N-terminal end and detected protein expression using IFA and Western blotting. Tagging the protein with 6xHA did not significantly affect the development of male and female gametocytes , ookinete conversion, the numbers of oocysts or SG sporozoites. PyAP2-G3 is strongly expressed in the cytoplasm and to a lesser degree in the nuclei of asexual stages and gametocytes (all the blood stages), but not in ookinetes. |
Gametocyte/Gamete | We tagged PyAP2-G3 with 6xHA at the N-terminal end and detected protein expression using IFA and Western blotting. Tagging the protein with 6xHA did not significantly affect the development of male and female gametocytes , ookinete conversion, the numbers of oocysts or SG sporozoites. PyAP2-G3 is strongly expressed in the cytoplasm and to a lesser degree in the nuclei of asexual stages and gametocytes (all the blood stages), but not in ookinetes. |
Fertilization and ookinete | We tagged PyAP2-G3 with 6xHA at the N-terminal end and detected protein expression using IFA and Western blotting. Tagging the protein with 6xHA did not significantly affect the development of male and female gametocytes , ookinete conversion, the numbers of oocysts or SG sporozoites. PyAP2-G3 is strongly expressed in the cytoplasm and to a lesser degree in the nuclei of asexual stages and gametocytes (all the blood stages), but not in ookinetes. |
Oocyst | Not tested |
Sporozoite | Not tested |
Liver stage | Not tested |
Additional remarks phenotype | Mutant/mutation Other mutants |
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Details of the target gene | |||||||||||||||||||||||||||
Gene Model of Rodent Parasite | PY17X_1417400 | ||||||||||||||||||||||||||
Gene Model P. falciparum ortholog | PF3D7_1317200 | ||||||||||||||||||||||||||
Gene product | AP2 domain transcription factor, putative | ||||||||||||||||||||||||||
Gene product: Alternative name | ApiAP2; AP2-G3 | ||||||||||||||||||||||||||
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Details of the genetic modification | |||||||||||||||||||||||||||
Name of the tag | HA | ||||||||||||||||||||||||||
Details of tagging | 6xHA | ||||||||||||||||||||||||||
Additional remarks: tagging | To construct vectors for tagging PyApiAP2 genes with sequences coding for mCherry or 6xHA, we first amplified the C- or N-terminal part (300 to 800 bp) of the coding region as the left or right arm and 400 to 800 bp from the 5= UTR or 3' UTR following the translation stop codon as the right or left arm. A DNA fragment encoding the mCherry or 6xHA tag was inserted between the left and right arms in frame with the gene of interest. For each gene, one sgRNA was designed to target the site close to the C- or N-terminal part of the coding region. (see mutant RMgm-1096 for details). | ||||||||||||||||||||||||||
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 | ||||||||||||||||||||||||||
Plasmid/construct map | |||||||||||||||||||||||||||
Plasmid/construct sequence | |||||||||||||||||||||||||||
Restriction sites to linearize plasmid | |||||||||||||||||||||||||||
Selectable marker used to select the mutant parasite | hdhfr | ||||||||||||||||||||||||||
Promoter of the selectable marker | eef1a | ||||||||||||||||||||||||||
Selection (positive) procedure | pyrimethamine | ||||||||||||||||||||||||||
Selection (negative) procedure | No | ||||||||||||||||||||||||||
Additional remarks genetic modification | To construct vectors for tagging PyApiAP2 genes with sequences coding for mCherry or 6xHA, we first amplified the C- or N-terminal part (300 to 800 bp) of the coding region as the left or right arm and 400 to 800 bp from the 5= UTR or 3' UTR following the translation stop codon as the right or left arm. A DNA fragment encoding the mCherry or 6xHA tag was inserted between the left and right arms in frame with the gene of interest. For each gene, one sgRNA was designed to target the site close to the C- or N-terminal part of the coding region. (see mutant RMgm-1096 for details). | ||||||||||||||||||||||||||
Additional remarks selection procedure | |||||||||||||||||||||||||||
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