RMgmDB - Rodent Malaria genetically modified Parasites
SummaryRMgm-2151
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Last modified: 2 August 2017, 21:15
*RMgm-2151| Successful modification | The gene/parasite could not be changed/generated by the genetic modification. |
| The following genetic modifications were attempted | Gene disruption |
| Number of attempts to introduce the genetic modification | Unknown |
| Reference (PubMed-PMID number) |
Reference 1 (PMID number) : 28708996 |
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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 | Not applicable |
| Other information parent line | |
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| Attempts to generate the mutant parasite were performed by | |
| Name PI/Researcher | PlasmoGEM (O. Billker; J. Rayner) |
| Name Group/Department | PlasmoGEM |
| Name Institute | Wellcome Trust Sanger Institute |
| City | Hinxton Cambridge |
| Country | UK |
Disrupted: Mutant parasite with a disrupted gene| top of page | |||||||||||||||||||||||||
| Details of the target gene | |||||||||||||||||||||||||
| Gene Model of Rodent Parasite | PBANKA_0621100 | ||||||||||||||||||||||||
| Gene Model P. falciparum ortholog | PF3D7_0723600 | ||||||||||||||||||||||||
| Gene product | proteasome assembly chaperone 4, putative | ||||||||||||||||||||||||
| Gene product: Alternative name | |||||||||||||||||||||||||
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| Details of the genetic modification | |||||||||||||||||||||||||
| Inducable system used | No | ||||||||||||||||||||||||
| Additional remarks inducable system | |||||||||||||||||||||||||
| 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 | ||||||||||||||||||||||||
| Plasmid/construct map | |||||||||||||||||||||||||
| Plasmid/construct sequence | |||||||||||||||||||||||||
| Restriction sites to linearize plasmid | |||||||||||||||||||||||||
| Partial or complete disruption of the gene | Complete | ||||||||||||||||||||||||
| Additional remarks partial/complete disruption | |||||||||||||||||||||||||
| 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 | The gene is 'likely essential' for asexual blood stage growth/multiplication as determined by barcode PCR in a large pool of gene-deletion mutants. P. berghei blood stages were transfected with a large pool of barcoded disruption (gene-deletion) vectors. These disruption vectors contained long modification arms to efficiently target the genes of interest. In addition the vectors contain gene-specific molecular barcodes. Co-transfecting multiple gene-deletion vectors in the same electroporation reproducibly generates complex pools of barcoded P. berghei mutants. Unsuccessful gene disruption/deletion or successful gene disruption/deletion is determined by the 'absence or presence' of the barcode in the population (as determined by barcode-specific PCR analysis). This report is based on simultaneous phenotyping of mutants by barcode sequencing and is part of a large-scale genetic screen. Asexual blood stage phenotype: Growth rate phenotypes were obtained by counting barcodes on a next generation sequencer daily between days 4 and 8 post transfection. It is in the nature of the screen that genotypes of individual mutants were not validated. * Essential: growth rate not significantly different from 0.1. * Dispensable: growth rate not significantly different from 1 (corresponding to wild type). * Slow: growth rate significantly above 0.1 but below 1 (p < 0.05). * Fast: growth rate significantly above 1 (p<0.05). The project website http://plasmogem.sanger.ac.uk/ has further information on methods and vector designs, provides tools for data visualisation and analysis, and allows researchers to request vectors to recreate mutants, confirm individual phenotypes and conduct more in-depth analyses. | ||||||||||||||||||||||||
| Additional remarks selection procedure | |||||||||||||||||||||||||
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Primer information: Primers used for amplification of the target sequences
 Primer information: Primers used for amplification of the target sequences
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