Back to search resultsSummaryRMgm-752
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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) : 20227667 Reference 2 (PMID number) : 22426943 |
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 | A reference wild type clone from the ANKA strain of P. berghei (PubMed: PMID: 17406255) |
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The mutant parasite was generated by | |
Name PI/Researcher | S.Iwanaga; C.J. Janse; A.P. Waters; M. Yuda |
Name Group/Department | School of Medicine |
Name Institute | Mie University |
City | Tsu City |
Country | Japan |
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Name of the mutant parasite | |
RMgm number | RMgm-752 |
Principal name | C-PAC; L-PAC |
Alternative name | |
Standardized name | |
Is the mutant parasite cloned after genetic modification | Yes |
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Phenotype | |
Asexual blood stage | Stable segregation and maintenance of circular and linear 'Plasmodium artificial chromosomes' (C-PAC; L-PAC). |
Gametocyte/Gamete | Stable segregation and maintenance of circular and linear 'Plasmodium artificial chromosomes' (C-PAC; L-PAC). |
Fertilization and ookinete | Stable segregation and maintenance of circular and linear 'Plasmodium artificial chromosomes' (C-PAC; L-PAC). |
Oocyst | Stable segregation and maintenance of circular and linear 'Plasmodium artificial chromosomes' (C-PAC; L-PAC). |
Sporozoite | Stable segregation and maintenance of circular and linear 'Plasmodium artificial chromosomes' (C-PAC; L-PAC). |
Liver stage | Stable segregation and maintenance of circular and linear 'Plasmodium artificial chromosomes' (C-PAC; L-PAC). |
Additional remarks phenotype | Mutant/mutation The C-PAC is a circular construct containing a Plasmodium centromere. The L-PAC is a linear construct containing a Plasmodium centromere and Plasmodium telomeric sequences See the following papers for more information: Iwanaga S, Khan SM, Kaneko I, Christodoulou Z, Newbold C, Yuda M, Janse CJ, Waters AP. Functional identification of the Plasmodium centromere and generation of a Plasmodium artificial chromosome. Cell Host Microbe. 2010 Mar 18;7(3):245-55. |
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Details of the target gene | |
Gene Model of Rodent Parasite | Not available |
Gene Model P. falciparum ortholog | Not available |
Gene product | Not available |
Gene product: Alternative name | |
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Description | |
Short description of the modification | Mutants containing circular or linear 'Plasmodium artificial chromosomes' (C-PAC; L-PAC). |
Description | To generate a Plasmodium artificial chromosome (PAC), we had to first functionally identify and characterize the parasite's centromere. A putative centromere (pbcen5) was cloned from chromosome 5 of the rodent parasite P. berghei based on a Plasmodium gene-synteny map. Plasmids containing pbcen5 were stably maintained in parasites during a blood-stage infection with high segregation efficiency, without drug pressure. pbcen5-containing plasmids were also stably maintained during parasite meiosis and mitosis in the mosquito. A linear PAC (L-PAC) was generated by integrating pbcen5 and telomere into a plasmid. The L-PAC segregated with a high efficiency and was stably maintained throughout the parasite's life cycle, as either one or two copies. These results suggest that L-PAC behaves like a Plasmodium chromosome, which can be exploited as an experimental research tool. Construction of Plasmids Containing PCEN (C-PAC) The various (complete and partial) PCEN-containing constructs were cloned into the plasmid vector pbGFPcon, which contains a selectable marker cassette encoding the Toxoplasma gondii pyrimethamine-resistant dhfr-ts gene (Tgdhfr-ts) as well as a gfp expression cassette under the control of the P. berghei eef1αa promoter. pbGFPcon was digested with EcoRI, thereby removing its d-ssu-rrna sequence. Subsequently, the PCEN-containing DNA fragments were digested with EcoRI and cloned into the EcoRI-digested pbGFPcon. Construction of the Plasmodium Artificial Chromosome (L-PAC) The DNA fragment containing the two telomeric sequences, oriented head to head with an ∼500 bp spacer region, was digested with HindIII and then cloned into the HindIII-digested pbCEN5A/T plasmid. This resulted in a circular plasmid, termed C-PAC. When we digested the C-PAC with PmeI, removing the spacer region between the two telomeres, the remaining linearized fragment was designated L-PAC. |
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