Summary

RMgm-495
Malaria parasiteP. berghei
Genotype
DisruptedGene model (rodent): PBANKA_1033100; Gene model (P.falciparum): PF3D7_1409400; Gene product: conserved Plasmodium membrane protein, unknown function
PhenotypeNo phenotype has been described
Last modified: 21 December 2011, 09:49
  *RMgm-495
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 676m1cl1 (RMgm-29)
Other information parent line676m1cl1 (RMgm-29) is a reference ANKA mutant line which expresses GFP-luciferase under control of a constitutive promoter (PubMed: PMID: 16242190).
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-495
Principal nameexp. 1499
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_1033100
Gene Model P. falciparum ortholog PF3D7_1409400
Gene productconserved Plasmodium membrane 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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GTACCCCCATATTTATTTTTTGCTATTCAAGAATAACTTGAAATAAAAAATAAATATATT
TATTATACATATAAAAGACTCAAGAAAACATAAAATACCTAATTAAATATAAAACAAAAT
GGATTCAGGTATAACGAAACATAAATATTTCATAAATGAGCTTACACATATATAAATGCT
TAAATGGTCTTTGCTTATACATGAACAATAATGGGGGGGGATGCATATACGCAGATACAT
ACATATATATACATTATATTCATTGGTATACACAAACGCGTATGACACAACTGCACACAT
ACGCGTATACACACATAAATGTGATAATGTGTGTTTTGTGGCGAAAGAAAAACCTTAAGT
GTGCATTTTAGAGATTTTATACTATCAAATTTATATATGTTATATATGCATATGATACAC
GCAATTATTTGTGGTGAAATACAAATATCAGGCATTTTATAAATTTATTTAGATATTCAT
GTTAACGGTAATTACTTTGACATTATTATATAAATTGTGATATATATTTTTTTATATAGA
ATATCAAGGTTTATTAAACGGTAAAGAAAAGGAAGATGAAACAAGCTTGCATGCCTGCAG
GTCAACAATAAATAATAAATAAATATTGTGGAAATAAAATAACATATAATTATTTTTAAT
ACATTGATTTCCCTTTTATTTTTTTAAATTTCATTGATATAAAAATATATAATAATAACA
TATATGATTTCAAATTAATCTTTTCAAAAATGGTGTTTATTTTTGTATGTTTGTGTATGA
ATTAATCACATAACACATCTATTAAATTGAGTTGGTAATATAGACACAAATAAATATATA
TATTTTTATAGCTTAAAAGTGTGTTATGAATATTTTAAGCATATTTTCTTTTTCTTTGGA
TTGTGTAAAATGAACTCATATAATGCGTTTTTTTGTTTTTGTTATTTTGTCATTTTGTTA
TTTTGCTATTTTATGGATTAATTTTTGTTTATAAAATGGGAAATAATTTTAACATATTTA
AATAAATGGAGAAAAAATATAAAATAATTATAAAAAAAAGTTAATACACATTTTTTCCTG
TTATAGACCTTATATTTATTTATCCATATATATATATATATATATATATATATATATATA
CATACCAAGTGAATTAAGAGGAAAGCTAATTTATTATTCAGAATAATATATGAACTATAT
ATAATTTTTATTATTTTGGTGTATATTAATCTGTCTATATGCATACATGCAATAATTTAT
CGACTTATATATCAAATAACATAAAATAGAAGTGTTTTAAATTATGGATATATGCTCAAT
ATTCATTTTTTTTAATAAGTTAGCTATATTTAAATTATACATTTTATATATGGTCTCTTT
TTTTTTTAAATATTATTTAAGTGATCATGAAAATATAAATAATTTTTTTTTATTTAATAT
CCTTTTGCTTGCATGTGGTAAATGGAAATTTGGATGTGTTTTGAARGTTCGGATATAGTT
GTATGGACATATAATATATTTTGTGAAAAATTGGTTTTATGTTTATACTTATGCCAATAC
TTTTTGAGTAAAACAAAGCAAGTGCTTATAAATAATTAAAGCCAATTTTATAATATATAT
TTTTTTATTTAATTTGAATTTAGTAGTATAATTTTTTATGGTAAGTGCTCAAAGAGAGTT
GCTTATAAAGTATGGTTTGTTTCTTTTTCGCCATTTTGAATTACACATTAAAAATATATA
GATACATATATTATAATATGAAATCATTAATAATTTAGGGAAATTCTACAAATTTAAAAA
CGAATAAAATAATTGTTTTTCATCATGCCATAACACAATATTGATATATACATGTACAAA
CATTTTTTTTATTTGGAAAATATAAATTATATAAAAAAAAATGTATAGTATACAAAATGA
GCATATTCACACGGGGTGGACGTTCATTTTTTCATTTTTCCCCTGTTTTTTATGAGTATA
TGATAAAATTTTATGAACATTTACACAAAATGAAAATGGATATATAGGAAAAATGGAGCG
GTATTTCATTTATCTTTGATTGTCATTTGGATATTATATTACCYTGGGTAGGCAATTAAA
AATGTTAAATAACAATTTAAGGAAATTATATTTTATATATTAAAATTAACACTGTATTAT
ATGATTCGCTTATAAAAGCCACTCTTTCCCCATGCAAAGCTGTTTAATATCAATTTTAAC
AAATTACACACATGTTAATATATTTATATATATAATTTATATATTTATAATTTATATATT
TATATTTTTATTATTTATATATTTATTATTTATTGTGTGTGTCAATTCGGGTAGGATATA
CCTCTTTTTTATTGTTTAAAGCGATTTGTATTCTAAAATATAAAGRATTTGAAAAAGAGA
AAGATAGAATATGATCCCATCATATATAGCCCTATAATTTTTATTTAGCAGCGAATTAAT
TTTTCTATTAAGTTTATGTGTAATTAAAATAACGGAATATATATAATACAATAAAAAAGT
GCATAAATTAAAATTTTTTCAATTAAATTTTTTTTTTTAAGGGGTTATATAATATTAAAT
ATATAAAATACGATTATATATTTTTGCTACAATTTTTTATATTAAGATATAAATAGTAAA
TAAATGGTATTATATGGCATGTAATATATAAATTTTTTCCAATTTTTATTTTATATACAC
TTTTCCTTTTTTTGTCATAAAACTTAAACAATTTACACATTCATTTTAAAAATTGACTAT
TTGTTTCAACATTTTTTGAGTTTCCGTTTTATAATAGTATTTTCATTTGTATATTGCTTA
TATATATAAATACACACCTAAATGTTACAAAGGATCAATGCATAAACCGGTGTGTCTGGT
CGTCGCGATGACCCCCAAGAGGGGCATCGGCATCAACAACGGCCTCCCGTGGCCCCACTT
GACCACAGATTTCAAACACTTTTCTCGTGTGACAAAAACGACGCCCGAAGAAGCCAGTCG
CCTGAACGGGTGGCTTCCCAGGAAATTTGCAAAGACGGGCGACTCTGGACTTCCCTCTCC
ATCAGTCGGCAAGAGATTCAACGCCGTTGTCATGGGACGGAAAACCTGGGAAAGCATGCC
TCGAAAGTTTAGACCCCTCGTGGACAGATTGAACATCGTCGTTTCCTCTTCCCTCAAAGA
AGAAGACATTGCGGCGGAGAAGCCTCAAGCTGAAGGCCAGCAGCGCGTCCGAGTCTGTGC
TTCACTCCCAGCAGCTCTCAGCCTTCTGGAGGAAGAGTACAAGGATTCTGTCGACCAGAT
TTTTGTCGTGGGAGGAGCGGGACTGTACGAGGCAGCGCTGTCTCTGGGCGTTGCCTCTCA
CCTGTACATCACGCGTGTAGCCCGCGAGTTTCCGTGCGACGTTTTCTTCCCTGCGTTCCC
CGGAGATGACATTCTTTCAAACAAATCAACTGCTGCGCAGGCTGCAGCTCCTGCCGAGTC
TGTGTTCGTTCCCTTTTGTCCGGAGCTCGGAAGAGAGAAGGACAATGAAGCGACGTATCG
ACCCATCTTCATTTCCAAGACCTTCTCAGACAACGGGGTTCCCTACGACTTTGTGGTTCT
CGAGAAGAGAAGGAAGACTGACGACGCAGCCACTGCGGAACCGAGCAACGCAATGAGCTC
CTTGACGTCCACGAGGGAGACAACTCCCGTGCACGGGTTGCAGGCTCCTTCTTCGGCCGC
AGCCATTGCCCCGGTGTTGGCGTGGATGGACGAAGAAGACCGGAAAAAACGCGAGCAAAA
GGAACTGATTCGGGCCGTTCCGCATGTTCACTTTAGAGGCCATGAAGAGTTCCAGTACCT
TGATCTCATTGCCGACATTATTAACAATGGAAGGACAATGGATGACCGAACGGGCGTTGG
TGTCATCTCCAAATTCGGCTGCACTATGCGCTACTCGCTGGATCAGGCCTTTCCACTTCT
CACCACAAAGCGTGTGTTCTGGAAAGGGGTCCTCGAAGAGTTGCTGTGGTTCATTCGCGG
CGACACGAACGCAAACCATCTTTCTGAGAAGGGCGTGAAGATCTGGGACAAGAATGTGAC
ACGCGAGTTCCTCGATTCGCGCAATCTCCCCCACCGAGAGGTCGGAGACATCGGCCCGGG
CTACGGCTTCCAGTGGAGACACTTCGGCGCGGCATACAAAGACATGCACACAGACTACAC
AGGGCAGGGCGTCGACCAGCTGAAGAATGTGATCCAGATGCTGAGAACGAATCCAACAGA
TCGTCGCATGCTCATGACTGCCTGGAATCCTGCAGCGCTGGACGAAATGGCGCTGCCGCC
TTGTCACTTGTTGTGCCAGTTCTACGTGAACGACCAGAAGGAGCTGTCGTGCATCATGTA
TCAGCGGTCGTGCGATGTCGGCCTCGGCGTCCCCTTCAACATCGCTTCCTATTCGCTTTT
GACGCTCATGGTTGCACACGTCTGCAACCTAAAACCTAAGGAGTTCATTCACTTCATGGG
GAACACGCATGTCTACACGAACCATGTCGAGGCTTTAAAAGAGCAGCTGCGGAGAGAACC
GAGACCGTTCCCCATTGTGAACATCCTCAACAAGGAACGCATCAAGGAAATCGACGATTT
CACCGCCGAGGATTTTGAGGTCGTGGGCTACGTCCCGCACGGACGAATCCAGATGGAGAT
GGCTGTCTAGCGGAAATACAGAAGCTAGCTTTGATCCCGTTTTTCTTACTTATATATTTA
TACCAATTGATTGTATTTATAACTGTAAAAATGTGTATGTTGTGTGCATATTTTTTTTTG
TGCATGCACATGCATGTAAATAGCTAAAATTATGAACATTTTATTTTTTGTTCAGAAAAA
AAAAACTTTACACACATAAAATGGCTAGTATGAATAGCCATATTTTATATAAATTAAATC
CTATGAATTTATGACCATATTAAAAATTTAGATATTTATGGAACATAATATGTTTGAAAC
AATAAGACAAAATTATTATTATTATTATTATTTTTACTGTTATAATTATGTTGTCTCTTC
AATGATTCATAAATAGTTGGACTTGATTTTTAAAATGTTTATAATATGATTAGCATAGTT
AAATAAAAAAAGTTGAAAAATTAAAAAAAAACATATAAACACAAATGATGTTTTTTCCTT
CAATTTCGATATCGAATTCGAATGGATCAACGGTCCAGTTACATACTTGTAAAATATATT
TATGCAAGTATCGTTGTGCATGCTTACATATATAATAGACATTTTTATAGTTATTTACTT
CTACATCCATTATTATACAGTTATTAAATAATAGGAAGAGCAAGATCAATGTGCTTAGTT
ATTTTGTTTCATTATTTATATTTTTTATTTATTTTTTTGTGGTCTATTAAGTTTTTGTTT
ATATTATGAAAAAATTTTTATTAACTTAATTTTTTTCATTATTATTCCCTTTTTAACACT
AAAATTCATAAGTGAAAAGTTGTCACAAAAAAGTATTTTTTCTTTTTAAATATTTTATTT
AATTTATTATGGTTATTATTATCTTAAGTTTGTGACCATACATTATTGTAAAAGATTTCA
AACAATATTTTTTAAATCAAGGTACTGTCTTTTTGGGGAAAGTGACATATCCTTGGAACA
AGAAAATCCCTTCATATAAGTATGTGCACTAATTTAGTATATATATGATCTTTGTGGTCT
AACATGAAATAGATGTATCCATGTTAATACGATAAGGGAAACAAAGTATATATACAATGT
AATTTGAATGCTCTTATACGATTTTATAATAATATAAAACTATAATGCGAATACTTTTAT
TTATTATTATATTTATACATATGTCTATACAGTAATTTATACACATTTTTTGTCAAAGAA
CATGGCATTTTCCCTCATACACTCTAGAGCGGCCGCCACCGCGGTGGAGCTCCAATTCGC
CCTATAGTGAGTCGTATTACAATTCACTGGCCGTCGTTTTACAACGTCGTGACTGGGAAA
ACCCTGGCGTTACCCAACTTAATCGCCTTGCAGCACATCCCCCTTTCGCCAGCTGGCGTA
ATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGTTGCGCAGCCTGAATGGCGAAT
GGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGTGGTGGTTACGCGCAGCGTGA
CCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGCTTTCTTCCCTTCCTTTCTCG
CCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGGCTCCCTTTAGGGTTCCGAT
TTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGGTGATGGTTCACGTAGTG
GGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAGTCCACGTTCTTTAATA
GTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGTCTATTCTTTTGATT
TATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTAACAAAAAT
TTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTAGGTGGCACTTTTCGGGGAAA
TGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCAT
GAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCA
ACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCA
CCCAGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTA
CATCGAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTT
TCCAATGATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGC
CGGGCAAGAGCAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTC
ACCAGTCACAGAAAAGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGC
CATAACCATGAGTGATAACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAA
GGAGCTAACCGCTTTTTTGCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGA
ACCGGAGCTGAATGAAGCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAAT
GGCAACAACGTTGCGCAAACTATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACA
ATTAATAGACTGGATGGAGGCGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCC
GGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCAT
TGCAGCACTGGGGCCAGATGGTAAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAG
TCAGGCAACTATGGATGAACGAAATAGACAGATCGCTGAGATAGGTGCCTCACTGATTAA
GCATTGGTAACTGTCAGACCAAGTTTACTCATATATACTTTAGATTGATTTAAAACTTCA
TTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGATAATCTCATGACCAAAATCCC
TTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAGAAAAGATCAAAGGATCTTC
TTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAAACCACCGCTACC
AGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCGAAGGTAACTGGCTT
CAGCAGAGCGCAGATACCAAATACTGTCCTTCTAGTGTAGCCGTAGTTAGGCCACCACTT
CAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGC
TGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTTACCGGATAA
GGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGGAGCGAACGAC
CTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCTTCCCGAAGG
GAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGCGCACGAGGGA
GCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCACCTCTGACT
TGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAACGCCAGCAA
CGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGTTCTTTCCTGC
GTTATCCCCTGATTCTGTGGATAACCGTATTACCGCCTTTGAGTGAGCTGATACCGCTCG
CCGCAGCCGAACGACCGAGCGCAGCGAGTCAGTGAGCGAGGAAGCGGAAGAGCGCCCAAT
ACGCAAACCGCCTCTCCCCGCGCGTTGGCCGATTCATTAATGCAGCTGGCACGACAGGTT
TCCCGACTGGAAAGCGGGCAGTGAGCGCAACGCAATTAATGTGAGTTAGCTCACTCATTA
GGCACCCCAGGCTTTACACTTTATGCTTCCGGCTCGTATGTTGTGTGGAATTGTGAGCGG
ATAACAATTTCACACAGGAAACAGCTATGACCATGATTACGCCAAGCTCGAAATTAACCC
TCACTAAAGGGAACAAAAGCTG
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 1GCCGGTACCCCCATATTTATTTTTTGCTATTC
Additional information primer 1F1 primer
Sequence Primer 2GCCAAGCTTGTTTCATCTTCCTTTTCTTTACCG
Additional information primer 2R1 primer
Sequence Primer 3GCCGAATTCGAATGGATCAACGGTCCAGTTAC
Additional information primer 3F2 primer
Sequence Primer 4GCCTCTAGAGTGTATGAGGGAAAATGCCATGTTC
Additional information primer 4R2 primer
Sequence Primer 5
Additional information primer 5
Sequence Primer 6
Additional information primer 6