SummaryRMgm-402
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Successful modification | The parasite was generated by the genetic modification |
The mutant contains the following genetic modification(s) | Other |
Reference (PubMed-PMID number) |
Reference 1 (PMID number) : 21418605 |
MR4 number | |
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Parent parasite used to introduce the genetic modification | |
Rodent Malaria Parasite | P. berghei |
Parent strain/line | P. berghei ANKA |
Name parent line/clone | P. berghei ANKA cl15cy1 |
Other information parent line | |
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The mutant parasite was generated by | |
Name PI/Researcher | J. Fonager; C.J. Janse; A.P. Waters |
Name Group/Department | Leiden Malara Research Group |
Name Institute | Leiden University Medical Center |
City | Leiden |
Country | The Netherlands |
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Name of the mutant parasite | |
RMgm number | RMgm-402 |
Principal name | piggyBac P5 (5) |
Alternative name | |
Standardized name | |
Is the mutant parasite cloned after genetic modification | No |
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Phenotype | |
Asexual blood stage | Not tested |
Gametocyte/Gamete | Not tested |
Fertilization and ookinete | Not tested |
Oocyst | Not tested |
Sporozoite | Not tested |
Liver stage | Not tested |
Additional remarks phenotype | Mutant/mutation A transposase mediated piggyBac insertion into the open reading frame (ORF). PiggyBac insertion into the TTAA tetranucleotide insertion site acgtaataattttttcttatttaatatttttattgcaactattt The piggyBac insert was identified using a TAIL PCR approach. The piggyBac insertion is expected to affect/disrupt expression of the gene. The presence of an insert in the ORF may therefore provide indirect evidence that the gene is NOT essential for blood stage development. However, neither expression or the phenotype of the mutant containing the insert have been analysed. See RMgm-521 for an independent mutant lacking expression of PBANKA_131800. This mutant has been generated by standard methods of gene disruption. Phenotype analyses indicate normal blood stage development. The mutant shows a strongly reduced sporozoite production. PiggyBac-mediated insertion was achieved when parasites containing the transposase gene stably integrated into the genome (see RMgm-406) were transfected with piggyBac donor plasmids. In the paper describing this mutant two different approaches were used to obtain insertion of piggyBac elements into the genome of P. berghei. In the first approach parasites were ‘co-transfected’ with both piggyBac donor plasmid (pL1302) and the helper plasmid (pL1301). The helper plasmid contains the transposase under the control of the constitutive eef1a promoter and this plasmid does not contain a drug-selection cassette (see Figure). Since plasmids are not retained in parasites during asexual growth without drug-selection the helper plasmid is ‘transiently transfected’ and will be lost from the parasites during blood stage growth. In the second approach the donor plasmid was transfected into transgenic parasites that contained the transposase gene stably integrated into the c/d-ssu-rrna gene locus. This transgenic line, TPSama1 (RMgm-406), was generated using standard methods for transfection of P. berghei and the transgenic parasites contain the T. gondii dhfr/ts as a selectable marker and transposase under the control of the schizont specific ama-1 promoter (see Figure). |
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Details of the target gene | |
Gene Model of Rodent Parasite | PBANKA_1318000 |
Gene Model P. falciparum ortholog | PF3D7_1454300 |
Gene product | SNF1-related serine/threonine protein kinase KIN |
Gene product: Alternative name | |
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Description | |
Short description of the modification | Transposase mediated piggyBac insertion into the ORF |
Description | See Additional Remarks Phenotype |
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