RMgmDB - Rodent Malaria genetically modified Parasites

Summary

RMgm-4405

Malaria parasite	P. yoelii
Genotype
Disrupted	Gene model (rodent): PY17X_1434600; Gene model (P.falciparum): PF3D7_1216600; Gene product: cell traversal protein for ookinetes and sporozoites (CelTOS, S4)
Phenotype	Fertilization and ookinete; Oocyst; Sporozoite;

Last modified: 1 January 2018, 19:31

*RMgm-4405

Successful modification	The parasite was generated by the genetic modification
The mutant contains the following genetic modification(s)	Gene disruption
Reference (PubMed-PMID number)	Reference 1 (PMID number) : 29253313
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	Steel RWJ, Kappe SHI
Name Group/Department	Center for Infectious Disease Research
Name Institute	formerly Seattle Biomedical Research Institute
City	Seattle
Country	USA
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Name of the mutant parasite
RMgm number	RMgm-4405
Principal name	Py s4/celtos(-)
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modification	Yes
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Phenotype
Asexual blood stage	Not different from wild type
Gametocyte/Gamete	Not different from wild type
Fertilization and ookinete	Normal gametocyte and ookinete production. Surprisingly PPy s4/celtos¯ parasites also established a low-level mosquito midgut infection.
Oocyst	Normal gametocyte and ookinete production. Surprisingly Py s4/celtos¯ parasites also established a low-level mosquito midgut infection. Whereas the number of oocysts per midgut stayed stable between days 7 – 11 for Py WT, oocyst numbers for Py s4/celtos¯ parasites declined throughout this period. The fraction of midguts infected with Py s4/celtos¯ declined markedly from day 7 - 11, by which time oocysts were only occasionally observed and these exhibited defects in sporulation compared to Py WT oocysts. Combined, the data suggest that Py S4/CelTOS has an important role during oocyst survival and/or sporogenesis in addition to the previously reported role in ookinete traversal of the midgut epithelium.
Sporozoite	No salivary gland sporozoites.
Liver stage	Not tested
Additional remarks phenotype	Mutant/mutation The mutant lacks expression of CelTOS (S4) Protein (function) CelTOS is localized to the micronemes of ookinetes and (salivary gland) sporozoites and is secreted. It plays a role in migration of ookinetes through mosquito midgut cells and traversal of sporozoites through heptocytes Phenotype Normal gametocyte and ookinete production. Surprisingly Py s4/celtos¯ parasites also established a low-level mosquito midgut infection. Whereas the number of oocysts per midgut stayed stable between days 7 – 11 for Py WT, oocyst numbers for Py s4/celtos¯ parasites declined throughout this period. The fraction of midguts infected with Py s4/celtos¯ declined markedly from day 7 - 11, by which time oocysts were only occasionally observed and these exhibited defects in sporulation compared to Py WT oocysts. Combined, the data suggest that Py S4/CelTOS has an important role during oocyst survival and/or sporogenesis in addition to the previously reported role in ookinete traversal of the midgut epithelium. No salivary gland sporozoites. Additional information Other mutants

Disrupted: Mutant parasite with a disrupted gene

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Details of the target gene

Gene Model of Rodent Parasite

PY17X_1434600

Gene Model P. falciparum ortholog

PF3D7_1216600

Gene product

cell traversal protein for ookinetes and sporozoites

Gene product: Alternative name

CelTOS, S4

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Details of the genetic modification

Inducable system used

Additional remarks inducable system

Type of plasmid/construct used

(Linear) plasmid double cross-over

PlasmoGEM (Sanger) construct/vector used

Modified PlasmoGEM construct/vector used

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

Additional remarks genetic modification

To generate Py s4/celtos¯, two DNA fragments containing approximately 800 bp of the 5′ UTR and 3′ UTR of Py S4/CelTOS were amplified from WT Py 17XNL genomic DNA and cloned into the pDEF vector (BEI Resources).

Additional remarks selection procedure

Primer information: Primers used for amplification of the target sequences Click to view information

Primer information: Primers used for amplification of the target sequences Click to hide information

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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