Summary

RMgm-492
Malaria parasiteP. berghei
Genotype
DisruptedGene model (rodent): PBANKA_1038700; Gene model (P.falciparum): PF3D7_1403300; Gene product: conserved Plasmodium protein, unknown function
PhenotypeNo phenotype has been described
Last modified: 21 December 2011, 09:48
  *RMgm-492
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-492
Principal nameexp. 1488
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_1038700
Gene Model P. falciparum ortholog PF3D7_1403300
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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AGCTTGGGCCCCCGCGGTGGCGGCCGCTCTAGCTTTGATCCCGTTTTTCTTACTTATATA
TTTATACCAATTGATTGTATTTATAACTGTAAAAATGTGTATGTTGTGTGCATATTTTTT
TTTGTGCATGCACATGCATGTAAATAGCTAAAATTATGAACATTTTATTTTTTGTTCAGA
AAAAAAAAACTTTACACACATAAAATGGCTAGTATGAATAGCCATATTTTATATAAATTA
AATCCTATGAATTTATGACCATATTAAAAATTTAGATATTTATGGAACATAATATGTTTG
AAACAATAAGACAAAATTATTATTATTATTATTATTTTTACTGTTATAATTATGTTGTCT
CTTCAATGATTCATAAATAGTTGGACTTGATTTTTAAAATGTTTATAATATGATTAGCAT
AGTTAAATAAAAAAAGTTGAAAAATTAAAAAAAAACATATAAACACAAATGATGTTTTTT
CCTTCAATTTCGATTGATAATTCCTGCAGCCCAGCTTAATTCTTTTCGAGCTCTTTATGC
TTAAGTTTACAATTTAATATTCATACTTTAAGTATTTTTTGTAGTATCCTAGATATTGTG
CTTTAAATGCTCACCCCTCAAAGCACCAGTAATATTTTCATCCACTGAAATACCATTAAA
TTTTCAAAAAAATACTATGCATATAATGTTATACATATAAACATAAAACGCCATGTAAAT
CAAAAAATATATAAAAATATGTATAAAAATAAATATGCACTAAATATAAGCTAATTATGC
ATAAAAATTAAAGTGCCCTTTATTAACTAGAACTAGTCGTAATTATTTATATTTCTATGT
TATAAAAAAATCCTCATATAATAATATAATTAATATATGTAATGTTTTTTTTATTTTATA
ATTTTAATATAAAATAATATGTAAATTAATTCAAAAAATAAATATAATTGTTGTGAAACA
AAAAACGTAATTTTTTCATTTGCCTTCAAAATTTAAATTTATTTTAATATTTCCTAAAAT
ATATATACTTTGTGTATAAATATATAAAAATATATATTTGCTTATAAATAAATAAAAAAT
TTTATAAAACATAGGGGGATCCATGGTTGGTTCGCTAAACTGCATCGTCGCTGTGTCCCA
GAACATGGGCATCGGCAAGAACGGGGACCTGCCCTGGCCACCGCTCAGGAACGAATTTAG
ATATTTCCAGAGAATGACCACAACCTCTTCAGTAGAAGGTAAACAGAATCTGGTGATTAT
GGGTAAGAAGACCTGGTTCTCCATTCCTGAGAAGAATCGACCTTTAAAGGGTAGAATTAA
TTTAGTTCTCAGCAGAGAACTCAAGGAACCTCCACAAGGAGCTCATTTTCTTTCCAGAAG
TCTAGATGATGCCTTAAAACTTACTGAACAACCAGAATTAGCAAATAAAGTAGACATGGT
CTGGATAGTTGGTGGCAGTTCTGTTTATAAGGAAGCCATGAATCACCCAGGCCATCTTAA
ACTATTTGTGACAAGGATCATGCAAGACTTTGAAAGTGACACGTTTTTTCCAGAAATTGA
TTTGGAGAAATATAAACTTCTGCCAGAATACCCAGGTGTTCTCTCTGATGTCCAGGAGGA
GAAAGGCATTAAGTACAAATTTGAAGTATATGAGAAGAATGATGCTAGCGGAGGAGGTGG
ATCTGGTGGAGGTGGAAGTGCTAGCGTGACAGGGGGAATGGCAAGCAAGTGGGATCAGAA
GGGTATGGACATTGCCTATGAGGAGGCGGCCTTAGGTTACAAAGAGGGTGGTGTTCCTAT
TGGCGGATGTCTTATCAATAACAAAGACGGAAGTGTTCTCGGTCGTGGTCACAACATGAG
ATTTCAAAAGGGATCCGCCACACTACATGGTGAGATCTCCACTTTGGAAAACTGTGGGAG
ATTAGAGGGCAAAGTGTACAAAGATACCACTTTGTATACGACGCTGTCTCCATGCGACAT
GTGTACAGGTGCCATCATCATGTATGGTATTCCACGCTGTGTTGTCGGTGAGAACGTTAA
TTTCAAAAGTAAGGGCGAGAAATATTTACAAACTAGAGGTCACGAGGTTGTTGTTGTTGA
CGATGAGAGGTGTAAAAAGATCATGAAACAATTTATCGATGAAAGACCTCAGGATTGGTT
TGAAGATATTGGTGAGGCTTCGGAACCATTTAAGAACGTCTACTTGCTACCTCAAACAAA
CCAATTGCTGGGTTTGTACACCATCATCAGAAATAAGAATACAACTAGACCTGATTTCAT
TTTCTACTCCGATAGAATCATCAGATTGTTGGTTGAAGAAGGTTTGAACCATCTACCTGT
GCAAAAGCAAATTGTGGAAACTGACACCAACGAAAACTTCGAAGGTGTCTCATTCATGGG
TAAAATCTGTGGTGTTTCCATTGTCAGAGCTGGTGAATCGATGGAGCAAGGATTAAGAGA
CTGTTGTAGGTCTGTGCGTATCGGTAAAATTTTAATTCAAAGGGACGAGGAGACTGCTTT
ACCAAAGTTATTCTACGAAAAATTACCAGAGGATATATCTGAAAGGTATGTCTTCCTATT
AGACCCAATGCTGGCCACCGGTGGTAGTGCTATCATGGCTACAGAAGTCTTGATTAAGAG
AGGTGTTAAGCCAGAGAGAATTTACTTCTTAAACCTAATCTGTAGTAAGGAAGGGATTGA
AAAATACCATGCCGCCTTCCCAGAGGTCAGAATTGTTACTGGTGCCCTCGACAGAGGTCT
AGATGAAAACAAGTATCTAGTTCCAGGGTTGGGTGACTTTGGTGACAGATACTACTGTGT
TTAACTCGATCCCGTTTTTCTTACTTATATATTTATACCAATTGATTGTATTTATAACTG
TAAAAATGTGTATGTTGTGTGCATATTTTTTTTTGTGCATGCACATGCATGTAAATAGCT
AAAATTATGAACATTTTATTTTTTGTTCAGAAAAAAAAAACTTTACACACATAAAATGGC
TAGTATGAATAGCCATATTTTATATAAATTAAATCCTATGAATTTATGACCATATTAAAA
ATTTAGATATTTATGGAACATAATATGTTTGAAACAATAAGACAAAATTATTATTATTAT
TATTATTTTTACTGTTATAATTATGTTGTCTCTTCAATGATTCATAAATAGTTGGACTTG
ATTTTTAAAATGTTTATAATATGATTAGCATAGTTAAATAAAAAAAGTTGAAAAATTAAA
AAAAAACATATAAACACAAATGATGTTTTTTCCTTCAATTTCGGGTACGTACCCTCGAGG
CATGGTTTAGTCTAGGTAAATGTCTAGAGCGTAAGAACAATTATGATTTGTCTATTTTTT
CTTATAAAAATGCTATTTATTTTAGAAAGAATAACGTTTTCTATTATTTTTTGGCTAACA
TTTATTTTAAAACAGGACAAATAAAAAATTGCATAGGCATATTAAAATCTGCATGGAGGC
ACAATAAAAATTTTGTGTTTTCATCTATGTTGTTTGGTATATATTTAAAAAAGAAAAAAG
GGAATGTTTCTTTGCAAAACGAGGATCTAATTTTAGAAGAACATTGGAAAATAACTGATA
AATGCCATGTATGGTGTGTTAGATTTTTAAAAGATTATTTTAAAAAATTAAGAAACAAAT
ATTGTATGATGCAACTTTTTTGTGAGAACAAACGAGAGCAATATAATTTTTATGAGATGT
TATTATCACTTTTTAATTTGAACAAGCCTTCGATAAAGCATATACAATCAGTTAATTTTT
TGAGTTATTTTAATAAATATTATGAACAAAATTCAAGTTATTTTTATTTAATACGTACTT
TGAAGCATTCATCTTTAATTTTATACGACGGAATATATTATTTAGCCAATTATTTTCTTT
TTAATAAAAAAATAAAAAATGCTTTAATATTATTTAAAATTTTATGGGATGCAGGAGGAC
CTTATTCGGAATTCCTGCAGCCCGGGGGATCCACTAGTTCTAGAGCGGCCGCCACCGCGG
TGGAGCTCCAATTCGCCCTATAGTGAGTCGTATTACAATTCACTGGCCGTCGTTTTACAA
CGTCGTGACTGGGAAAACCCTGGCGTTACCCAACTTAATCGCCTTGCAGCACATCCCCCT
TTCGCCAGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGTTGCGC
AGCCTGAATGGCGAATGGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGTGGTG
GTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGCTTTC
TTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGGCTC
CCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGGT
GATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAG
TCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCG
GTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAG
CTGATTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTAGGTG
GCACTTTTCGGGGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAA
ATATGTATCCGCTCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGA
AGAGTATGAGTATTCAACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCC
TTCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGG
GTGCACGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTC
GCCCCGAAGAACGTTTTCCAATGATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTAT
TATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTCGCCGCATACACTATTCTCAGAATG
ACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTACGGATGGCATGACAGTAAGAG
AATTATGCAGTGCTGCCATAACCATGAGTGATAACACTGCGGCCAACTTACTTCTGACAA
CGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACATGGGGGATCATGTAACTC
GCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGACGAGCGTGACACCA
CGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATTAACTGGCGAACTACTTACTC
TAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGTTGCAGGACCACTTC
TGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCGGTGAGCGTG
GGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCCTCCCGTATCGTAGTTA
TCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACAGATCGCTGAGATAG
GTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATATATACTTTAGA
TTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGATAATC
TCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAGAAA
AGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAA
AAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTC
CGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTCCTTCTAGTGTAGCCGT
AGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCC
TGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGAC
GATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCA
GCTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCG
CCACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAG
GAGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGT
TTCGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTAT
GGAAAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTC
ACATGTTCTTTCCTGCGTTATCCCCTGATTCTGTGGATAACCGTATTACCGCCTTTGAGT
GAGCTGATACCGCTCGCCGCAGCCGAACGACCGAGCGCAGCGAGTCAGTGAGCGAGGAAG
CGGAAGAGCGCCCAATACGCAAACCGCCTCTCCCCGCGCGTTGGCCGATTCATTAATGCA
GCTGGCACGACAGGTTTCCCGACTGGAAAGCGGGCAGTGAGCGCAACGCAATTAATGTGA
GTTAGCTCACTCATTAGGCACCCCAGGCTTTACACTTTATGCTTCCGGCTCGTATGTTGT
GTGGAATTGTGAGCGGATAACAATTTCACACAGGAAACAGCTATGACCATGATTACGCCA
AGCTCGAAATTAACCCTCACTAAAGGGAACAAAAGCTGGTACCGTTTCGTATCTAGCAAT
TATGTGGAAATGCTATCATTCTTTTTATACATTTTTTTCTTTGTTTATTTTTATTTTATT
ATTACAATATTTATAATAGTCCGTTATTTTTTTAGAAATAAAAATACGAAGTGTCTTTAT
TTTTATTATGCTATTAAAATTTTTTTTAATTTAACTATATAAGTGATATTTTTAGTTTAT
TTAAAAAAACGAAAAAAGGATGAAATAATATAAGTAAAAATATGAAGGATATCCGAAGCT
GCAGTAATGTATCATTAAGAAGAAAACGGAAATATATCGAGGGTGAATCTAAAAAGTCCT
TGAATTTAATAAATAGACATAATAATGAAAATATTGTTAGTAAACCTAAGAGGAGATCAA
TAAGTTTGTCAAACGCATATTTTAGTGTAAACTCTTTTTCTATTTATGACTCATTAAATT
TTATGAATAATAATGGAATTAAGGGAGTGAAGAATGAAGAAGAAATTGGAAAAGTAGAAA
AGAAGGGGGAGGAAAACGACGCTAACGATAAAATAAGACAATATAAAAATAATGATATAC
GAAATAACAGTAATACGGAAAGA
Restriction sites to linearize plasmid KpnI/EcoRI
Partial or complete disruption of the geneUnknown
Additional remarks partial/complete disruption
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 1GCCGGTACCGTTTCGTATCTAGCAATTATGTG
Additional information primer 1F1 primer
Sequence Primer 2GCCAAGCTTCTTTCCGTATTACTGTTATTTCG
Additional information primer 2R1 primer
Sequence Primer 3GCCCTCGAGGCATGGTTTAGTCTAGGTAAATG
Additional information primer 3F2 primer
Sequence Primer 4GCCGAATTCCGAATAAGGTCCTCCTGCATCCC
Additional information primer 4R2 primer
Sequence Primer 5
Additional information primer 5
Sequence Primer 6
Additional information primer 6