Summary

RMgm-498
Malaria parasiteP. berghei
Genotype
DisruptedGene model (rodent): PBANKA_1331100; Gene model (P.falciparum): PF3D7_1467800; Gene product: conserved Plasmodium protein, unknown function
PhenotypeNo phenotype has been described
Last modified: 21 December 2011, 09:51
  *RMgm-498
Successful modificationThe 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) : 22184632
MR4 number
Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. berghei
Parent strain/lineP. berghei ANKA
Name parent line/clone P. berghei ANKA cl15cy1
Other information parent lineA reference wild type clone from the ANKA strain of P. berghei (PubMed: PMID: 17406255).
The mutant parasite was generated by
Name PI/ResearcherJ. Fonager, E.M. Pasini, C.J. Janse, B.M Franke-Fayard
Name Group/DepartmentLeiden Malaria Research Group
Name InstituteLeiden University Medical Center (LUMC)
CityLeiden
CountryThe Netherlands
Name of the mutant parasite
RMgm numberRMgm-498
Principal nameexp. 1560
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationNo
Phenotype
Asexual blood stageNot different from wild type
Gametocyte/GameteNot tested
Fertilization and ookineteNot tested
OocystNot tested
SporozoiteNot tested
Liver stageNot tested
Additional remarks phenotype

A pyrimethamine-resistant parasite population was selected after transfection with a construct for targeted gene disruption by double cross-over homologous recombination. The primers of the two target sequences of the gene are provided below.

The genotype of the parasites has not been analysed in detail.
Diagnostic PCR (for correct integration of the construct) and/or Southern analysis of PFG-separated chromosomes (to show integration into the chromosome on which the target gene is located) have been performed (see below). This analysis provided evidence for correct disruption of the gene indicating that it has no essential function during asexual blood stage development. However, no further analyses on the parasite population/cloned lines have been performed to show absence of transcripts or protein .

The phenotype of the selected parasite population/cloned lines has not been analysed in detail.
No obvious growth defects of the asexual blood stages of the selected parasite populations/cloned lines have been observed.

The gene has been selected for targeted gene disruption based on its presence in a proteome of membranes of infected red blood cells


  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_1331100
Gene Model P. falciparum ortholog PF3D7_1467800
Gene productconserved Plasmodium protein, unknown function
Gene product: Alternative name
Details of the genetic modification
Inducable system usedNo
Additional remarks inducable system
Type of plasmid/construct usedPlasmid double cross-over
PlasmoGEM (Sanger) construct/vector usedNo
Modified PlasmoGEM construct/vector usedNo
Plasmid/construct map
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Plasmid/construct sequence
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CCCCCTCGAGGTCGACGGTATCGATAAGCTTGCATGCCTGCAGGTCAACAATAAATAATA
AATAAATATTGTGGAAATAAAATAACATATAATTATTTTTAATACATTGATTTCCCTTTT
ATTTTTTTAAATTTCATTGATATAAAAATATATAATAATAACATATATGATTTCAAATTA
ATCTTTTCAAAAATGGTGTTTATTTTTGTATGTTTGTGTATGAATTAATCACATAACACA
TCTATTAAATTGAGTTGGTAATATAGACACAAATAAATATATATATTTTTATAGCTTAAA
AGTGTGTTATGAATATTTTAAGCATATTTTCTTTTTCTTTGGATTGTGTAAAATGAACTC
ATATAATGCGTTTTTTTGTTTTTGTTATTTTGTCATTTTGTTATTTTGCTATTTTATGGA
TTAATTTTTGTTTATAAAATGGGAAATAATTTTAACATATTTAAATAAATGGAGAAAAAA
TATAAAATAATTATAAAAAAAAGTTAATACACATTTTTTCCTGTTATAGACCTTATATTT
ATTTATCCATATATATATATATATATATATATATATATATATACATACCAAGTGAATTAA
GAGGAAAGCTAATTTATTATTCAGAATAATATATGAACTATATATAATTTTTATTATTTT
GGTGTATATTAATCTGTCTATATGCATACATGCAATAATTTATCGACTTATATATCAAAT
AACATAAAATAGAAGTGTTTTAAATTATGGATATATGCTCAATATTCATTTTTTTTAATA
AGTTAGCTATATTTAAATTATACATTTTATATATGGTCTCTTTTTTTTTTAAATATTATT
TAAGTGATCATGAAAATATAAATAATTTTTTTTTATTTAATATCCTTTTGCTTGCATGTG
GTAAATGGAAATTTGGATGTGTTTTGAARGTTCGGATATAGTTGTATGGACATATAATAT
ATTTTGTGAAAAATTGGTTTTATGTTTATACTTATGCCAATACTTTTTGAGTAAAACAAA
GCAAGTGCTTATAAATAATTAAAGCCAATTTTATAATATATATTTTTTTATTTAATTTGA
ATTTAGTAGTATAATTTTTTATGGTAAGTGCTCAAAGAGAGTTGCTTATAAAGTATGGTT
TGTTTCTTTTTCGCCATTTTGAATTACACATTAAAAATATATAGATACATATATTATAAT
ATGAAATCATTAATAATTTAGGGAAATTCTACAAATTTAAAAACGAATAAAATAATTGTT
TTTCATCATGCCATAACACAATATTGATATATACATGTACAAACATTTTTTTTATTTGGA
AAATATAAATTATATAAAAAAAAATGTATAGTATACAAAATGAGCATATTCACACGGGGT
GGACGTTCATTTTTTCATTTTTCCCCTGTTTTTTATGAGTATATGATAAAATTTTATGAA
CATTTACACAAAATGAAAATGGATATATAGGAAAAATGGAGCGGTATTTCATTTATCTTT
GATTGTCATTTGGATATTATATTACCYTGGGTAGGCAATTAAAAATGTTAAATAACAATT
TAAGGAAATTATATTTTATATATTAAAATTAACACTGTATTATATGATTCGCTTATAAAA
GCCACTCTTTCCCCATGCAAAGCTGTTTAATATCAATTTTAACAAATTACACACATGTTA
ATATATTTATATATATAATTTATATATTTATAATTTATATATTTATATTTTTATTATTTA
TATATTTATTATTTATTGTGTGTGTCAATTCGGGTAGGATATACCTCTTTTTTATTGTTT
AAAGCGATTTGTATTCTAAAATATAAAGRATTTGAAAAAGAGAAAGATAGAATATGATCC
CATCATATATAGCCCTATAATTTTTATTTAGCAGCGAATTAATTTTTCTATTAAGTTTAT
GTGTAATTAAAATAACGGAATATATATAATACAATAAAAAAGTGCATAAATTAAAATTTT
TTCAATTAAATTTTTTTTTTTAAGGGGTTATATAATATTAAATATATAAAATACGATTAT
ATATTTTTGCTACAATTTTTTATATTAAGATATAAATAGTAAATAAATGGTATTATATGG
CATGTAATATATAAATTTTTTCCAATTTTTATTTTATATACACTTTTCCTTTTTTTGTCA
TAAAACTTAAACAATTTACACATTCATTTTAAAAATTGACTATTTGTTTCAACATTTTTT
GAGTTTCCGTTTTATAATAGTATTTTCATTTGTATATTGCTTATATATATAAATACACAC
CTAAATGTTACAAAGGATCAATGCATAAACCGGTGTGTCTGGTCGTCGCGATGACCCCCA
AGAGGGGCATCGGCATCAACAACGGCCTCCCGTGGCCCCACTTGACCACAGATTTCAAAC
ACTTTTCTCGTGTGACAAAAACGACGCCCGAAGAAGCCAGTCGCCTGAACGGGTGGCTTC
CCAGGAAATTTGCAAAGACGGGCGACTCTGGACTTCCCTCTCCATCAGTCGGCAAGAGAT
TCAACGCCGTTGTCATGGGACGGAAAACCTGGGAAAGCATGCCTCGAAAGTTTAGACCCC
TCGTGGACAGATTGAACATCGTCGTTTCCTCTTCCCTCAAAGAAGAAGACATTGCGGCGG
AGAAGCCTCAAGCTGAAGGCCAGCAGCGCGTCCGAGTCTGTGCTTCACTCCCAGCAGCTC
TCAGCCTTCTGGAGGAAGAGTACAAGGATTCTGTCGACCAGATTTTTGTCGTGGGAGGAG
CGGGACTGTACGAGGCAGCGCTGTCTCTGGGCGTTGCCTCTCACCTGTACATCACGCGTG
TAGCCCGCGAGTTTCCGTGCGACGTTTTCTTCCCTGCGTTCCCCGGAGATGACATTCTTT
CAAACAAATCAACTGCTGCGCAGGCTGCAGCTCCTGCCGAGTCTGTGTTCGTTCCCTTTT
GTCCGGAGCTCGGAAGAGAGAAGGACAATGAAGCGACGTATCGACCCATCTTCATTTCCA
AGACCTTCTCAGACAACGGGGTTCCCTACGACTTTGTGGTTCTCGAGAAGAGAAGGAAGA
CTGACGACGCAGCCACTGCGGAACCGAGCAACGCAATGAGCTCCTTGACGTCCACGAGGG
AGACAACTCCCGTGCACGGGTTGCAGGCTCCTTCTTCGGCCGCAGCCATTGCCCCGGTGT
TGGCGTGGATGGACGAAGAAGACCGGAAAAAACGCGAGCAAAAGGAACTGATTCGGGCCG
TTCCGCATGTTCACTTTAGAGGCCATGAAGAGTTCCAGTACCTTGATCTCATTGCCGACA
TTATTAACAATGGAAGGACAATGGATGACCGAACGGGCGTTGGTGTCATCTCCAAATTCG
GCTGCACTATGCGCTACTCGCTGGATCAGGCCTTTCCACTTCTCACCACAAAGCGTGTGT
TCTGGAAAGGGGTCCTCGAAGAGTTGCTGTGGTTCATTCGCGGCGACACGAACGCAAACC
ATCTTTCTGAGAAGGGCGTGAAGATCTGGGACAAGAATGTGACACGCGAGTTCCTCGATT
CGCGCAATCTCCCCCACCGAGAGGTCGGAGACATCGGCCCGGGCTACGGCTTCCAGTGGA
GACACTTCGGCGCGGCATACAAAGACATGCACACAGACTACACAGGGCAGGGCGTCGACC
AGCTGAAGAATGTGATCCAGATGCTGAGAACGAATCCAACAGATCGTCGCATGCTCATGA
CTGCCTGGAATCCTGCAGCGCTGGACGAAATGGCGCTGCCGCCTTGTCACTTGTTGTGCC
AGTTCTACGTGAACGACCAGAAGGAGCTGTCGTGCATCATGTATCAGCGGTCGTGCGATG
TCGGCCTCGGCGTCCCCTTCAACATCGCTTCCTATTCGCTTTTGACGCTCATGGTTGCAC
ACGTCTGCAACCTAAAACCTAAGGAGTTCATTCACTTCATGGGGAACACGCATGTCTACA
CGAACCATGTCGAGGCTTTAAAAGAGCAGCTGCGGAGAGAACCGAGACCGTTCCCCATTG
TGAACATCCTCAACAAGGAACGCATCAAGGAAATCGACGATTTCACCGCCGAGGATTTTG
AGGTCGTGGGCTACGTCCCGCACGGACGAATCCAGATGGAGATGGCTGTCTAGCGGAAAT
ACAGAAGCTAGCTTTGATCCCGTTTTTCTTACTTATATATTTATACCAATTGATTGTATT
TATAACTGTAAAAATGTGTATGTTGTGTGCATATTTTTTTTTGTGCATGCACATGCATGT
AAATAGCTAAAATTATGAACATTTTATTTTTTGTTCAGAAAAAAAAAACTTTACACACAT
AAAATGGCTAGTATGAATAGCCATATTTTATATAAATTAAATCCTATGAATTTATGACCA
TATTAAAAATTTAGATATTTATGGAACATAATATGTTTGAAACAATAAGACAAAATTATT
ATTATTATTATTATTTTTACTGTTATAATTATGTTGTCTCTTCAATGATTCATAAATAGT
TGGACTTGATTTTTAAAATGTTTATAATATGATTAGCATAGTTAAATAAAAAAAGTTGAA
AAATTAAAAAAAAACATATAAACACAAATGATGTTTTTTCCTTCAATTTCGATATCGAAT
TCCTAATAACATCTTAATGGAATACATTTATTATTTTGAAATTTCATTAAATGTAATTTT
TTTTTTCCTCTTATAAAAAAAATGTTATATTTTATTATAACCACTTATTTTTTTAGTGCA
TACTATTCTTTAAGGAAAATACAAAAAAACTTTAAAAATATATAATTTGGTCTAAATATT
ATATATACATTTTTTATATTATTTCTTTTAGAATATATACATAAAAATTTTAAATATTTT
TTTTTTAATATAATTTGAGGAATATGAAAAGGACAACAAATTGTTATTTACCTATAAAAT
AAATAATATATATATGAATGTTTACCCAAATAAAAATTATAGTGCTAACTATATAAAAAA
AATATATTTTTTTCCATTAAAGAATTATTTCGATGTTAAAAATAAAAATATAATGCATAG
TTGGTATATTTTAATAATGTAACTTAATTTTTGGTGGATCTATAATTATAAAGGATATCC
TTCTCGTCATTTATATACCCTTTTTACAAAATTTATTTAAAAAAATTGTGAAAAAAAAAA
TATATTAACTTATTTTATATAGACAATGGTATATTTTTCTTAATTTTATGTAAGTCGCAG
TAGGTCATCAAAATAGTGAAAATAAAAAAGGGAAAATATTGATCACTTTGTTTTTTCATT
TTTTAAGCACACACAATCATTAAGCACGCGTATAAGCTCTAGAGCGGCCGCCACCGCGGT
GGAGCTCCAATTCGCCCTATAGTGAGTCGTATTACAATTCACTGGCCGTCGTTTTACAAC
GTCGTGACTGGGAAAACCCTGGCGTTACCCAACTTAATCGCCTTGCAGCACATCCCCCTT
TCGCCAGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGTTGCGCA
GCCTGAATGGCGAATGGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGTGGTGG
TTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGCTTTCT
TCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGGCTCC
CTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGGTG
ATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAGT
CCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGG
TCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGC
TGATTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTAGGTGG
CACTTTTCGGGGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAA
TATGTATCCGCTCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAA
GAGTATGAGTATTCAACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCT
TCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGG
TGCACGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCG
CCCCGAAGAACGTTTTCCAATGATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATT
ATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTCGCCGCATACACTATTCTCAGAATGA
CTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTACGGATGGCATGACAGTAAGAGA
ATTATGCAGTGCTGCCATAACCATGAGTGATAACACTGCGGCCAACTTACTTCTGACAAC
GATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACATGGGGGATCATGTAACTCG
CCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGACGAGCGTGACACCAC
GATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATTAACTGGCGAACTACTTACTCT
AGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGTTGCAGGACCACTTCT
GCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCGGTGAGCGTGG
GTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCCTCCCGTATCGTAGTTAT
CTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACAGATCGCTGAGATAGG
TGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATATATACTTTAGAT
TGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGATAATCT
CATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAGAAAA
GATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAA
AAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCC
GAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTCCTTCTAGTGTAGCCGTA
GTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCT
GTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACG
ATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAG
CTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGC
CACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGG
AGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTT
TCGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATG
GAAAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCA
CATGTTCTTTCCTGCGTTATCCCCTGATTCTGTGGATAACCGTATTACCGCCTTTGAGTG
AGCTGATACCGCTCGCCGCAGCCGAACGACCGAGCGCAGCGAGTCAGTGAGCGAGGAAGC
GGAAGAGCGCCCAATACGCAAACCGCCTCTCCCCGCGCGTTGGCCGATTCATTAATGCAG
CTGGCACGACAGGTTTCCCGACTGGAAAGCGGGCAGTGAGCGCAACGCAATTAATGTGAG
TTAGCTCACTCATTAGGCACCCCAGGCTTTACACTTTATGCTTCCGGCTCGTATGTTGTG
TGGAATTGTGAGCGGATAACAATTTCACACAGGAAACAGCTATGACCATGATTACGCCAA
GCTCGAAATTAACCCTCACTAAAGGGAACAAAAGCTGGTACCGAACAAAATATGAAATAA
ATTTGGATATAAAAACAGAAAATCATGAATATAATATTTCAACTTATTTTCCGCATTTTA
TTTCTGTTTTTTTTCATCAATACAATTTGTTTAAAATCTTAATTATACGAAAATATAATT
TTTTTTATTGGCCCGATATACTTGCCCTTTCATTAATAATATATATATTATTTTCTTTCT
TTTTTTTGAAAGCTTATTTTTTACATTATAATAACTTTATTAAAAAAAAAATTGATTGAT
ATACCTTGTATGAAAAATTTAACGATATTTTCCATTTTTTGAGAATATCTTTAACTTTAT
GATATAACTACATATACTGGATATATAAATAAATTACGCAAAAAAACAAAATGAGTGTGG
TATGTATATAAACAAAAAAAATGTAGATAAAAAAAGAAATTAAAATTTGTAAAAAATTTA
TATTTAAGCATATTAAATTATGATATTCCCATTAGTGTTACATTAATTTAAGCATATTTT
TATCAGTAATAATGTCATACTATTATATACACCCCATTTTGGAAACATTGAATATGTTAA
TAATTACACATTTTTCATTTTTTTCATCTCATTTAGAACACAATTACTGCTCGGGGCC
Restriction sites to linearize plasmid Asp718I/XbaI
Partial or complete disruption of the geneUnknown
Additional remarks partial/complete disruption
Selectable marker used to select the mutant parasitetgdhfr
Promoter of the selectable markerpbdhfr
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 1GCCGGTACCGAACAAAATATGAAATAAATTTGG
Additional information primer 1F1 primer
Sequence Primer 2GAGGGGGGGCCCCGAGCAGTAATTGTGTTCTAAATG
Additional information primer 2R1 primer
Sequence Primer 3GGGAATTCCTAATAACATCTTAATGGAATAC
Additional information primer 3F2 primer
Sequence Primer 4GGGAATTCCTAATAACATCTTAATGGAATAC
Additional information primer 4R2 primer
Sequence Primer 5
Additional information primer 5
Sequence Primer 6
Additional information primer 6