RMgmDB - Rodent Malaria genetically modified Parasites


Malaria parasiteP. berghei
Genetic modification not successful
DisruptedGene model (rodent): PBANKA_1450200; Gene model (P.falciparum): PF3D7_1235600; Gene product: Serine hydroxymethyltransferase, putative (SHMT)
PhenotypeNo phenotype has been described
Last modified: 9 August 2022, 17:11
Successful modificationThe 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 3
Reference (PubMed-PMID number) Reference 1 (PMID number) : 23173711
Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. berghei
Parent strain/lineP. berghei ANKA
Name parent line/clone Not applicable
Other information parent line
Attempts to generate the mutant parasite were performed by
Name PI/ResearcherPornthanakasem W, Leartsakulpanich U
Name Group/DepartmentNational Center for Genetic Engineering and Biotechnology
Name InstituteNational Center for Genetic Engineering and Biotechnology
CityKhlong Luang, Pathum Thani

  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_1450200
Gene Model P. falciparum ortholog PF3D7_1235600
Gene productSerine hydroxymethyltransferase, putative
Gene product: Alternative nameSHMT
Details of the genetic modification
Inducable system usedNo
Additional remarks inducable system
Type of plasmid/construct used(Linear) plasmid double cross-over
PlasmoGEM (Sanger) construct/vector usedNo
Modified PlasmoGEM construct/vector usedNo
Plasmid/construct map
Plasmid/construct sequence
Restriction sites to linearize plasmid
Partial or complete disruption of the geneComplete
Additional remarks partial/complete disruption The unsuccessful attempts to disrupt this gene indicate an essential function during asexual blood stage growth/multiplication.

Serine hydroxymethyltransferase (SHMT), a pyridoxal phosphate-dependent enzyme, plays a vital role in the de novo pyrimidine biosynthesis pathway in malaria parasites. Two genes have been identified in Plasmodium spp. encoding a cytosolic SHMT (cSHMT) and putative mitochondria SHMT (mSHMT).
Plasmodium falciparum contains PF3D7_1235600, a previously characterized cSHMT gene (Pfcshmt), and PF3D7_1456100, a putative gene of mSHMT (Pfmshmt).

Plasmids for the study of gene knockout in Plasmodium berghei ANKA strain were constructed based on the sequence of pL0017 vector (The Malaria Research and Reference Reagent Resource Center; MR4), which contains Toxoplasma gondii dihydrofolate reductase-thymidylate synthase (Tgdhfr/ts) and green fluorescent protein gene (gfp) expression cassettes for pyrimethamine (PYR) selection and fluorescence detection of transfected parasites. The 553 and 1,018 bp of PCR amplicons, corresponding to 5′- and 3′UTR of Pbcshmt (PBANKA_145020) respectively, were produced initially from P. berghei genomic DNA (gDNA). The 5′UTR fragment was inserted into pL0017 at Hind III site, while the 3′UTR fragment was inserted at Kpn I and Sac II sites respectively. This construct, pL0017_Δshmt, was used in the knockout study. For allelic replacement construct, gfp in pL0017_Δshmt was replaced with Plasmodium vivax cshmt (Pvcshmt; PVX_100730) and named pL0017_(Pv)Δshmt.
Selectable marker used to select the mutant parasitetgdhfr
Promoter of the selectable markereef1a
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modification
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