RMgmDB - Rodent Malaria genetically modified Parasites

Summary

RMgm-489
Malaria parasiteP. berghei
Genotype
DisruptedGene model (rodent): PBANKA_0836800; Gene model (P.falciparum): Not available; Gene product: erythrocyte membrane associated protein 1 fam-a protein (EMAP1: erythrocyte memberane associated protein 1)
PhenotypeNo phenotype has been described
Last modified: 18 January 2012, 12:41
  *RMgm-489
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
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Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. berghei
Parent strain/lineP. berghei ANKA
Name parent line/clone P. berghei ANKA 1037m1f1mocl1 (RMgm-32)
Other information parent lineP. berghei ANKA 1037m1f1mocl1 (1037cl1; RMgm-32) is a reference ANKA mutant line which expresses GFP-luciferase under control of a schizont-specific promoter (PubMed: PMID: 20019192). It does not contain a drug-selectable marker and has been selected by FACS sorting based on GFP expression
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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
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Name of the mutant parasite
RMgm numberRMgm-489
Principal nameexp. 1245
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
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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

This mutant (and this Pb-Fam-1 protein; named EMAP1) has been characterized in more detail in follow-up studies in the Leiden Malaria Research Group. These analyses will be published shortly.

  Disrupted: Mutant parasite with a disrupted gene
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Details of the target gene
Gene Model of Rodent Parasite PBANKA_0836800
Gene Model P. falciparum ortholog Not available
Gene producterythrocyte membrane associated protein 1 fam-a protein
Gene product: Alternative nameEMAP1: erythrocyte memberane associated protein 1
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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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ATCGCGACAGTGTTCGTGTTACATACGTCAACTCTGTAAGCAATATAAAAATTATAATAA
CATAATAATTTTATTTTATCATTATCAAAGATATATACCTTTTCTATAATTTGCCATATT
TTAATATTTCTATTCATCTACAAACAAATTGACCTATTTTATATTTCTATTCATTCTTTT
CTTTTTCCGTAGATTCATAATGGTGACTCCTTTCCAAAAGATTTTTGTGACAAACAATTT
AGAGTAAAAAATATGCTATATTTGATCGATTTATTAAAAAATAGTTTTAAATAATAAACG
TATACCCCACATAATGTTCTTTATCACTTTAATATTGAAATACCATATTAATAAAATTAA
ATTACTTTCATACACCATGCTTATTTTAAATTGCCATCACAAAACATCTTTTTTTGTTTT
ATGAAAGTTTATATCATATAATTTATTCACCCTTTTTAATATAATGGCAGCTCTACATGT
ATTTGATTATGCTTAGCACTATAAATGTATCCGAATTCCTGCAGCCCGGGGGATCCACTA
GTTCTAGAGCGGCCGCCACCGCGGTGGAGCTCCAATTCGCCCTATAGTGAGTCGTATTAC
AATTCACTGGCCGTCGTTTTACAACGTCGTGACTGGGAAAACCCTGGCGTTACCCAACTT
AATCGCCTTGCAGCACATCCCCCTTTCGCCAGCTGGCGTAATAGCGAAGAGGCCCGCACC
GATCGCCCTTCCCAACAGTTGCGCAGCCTGAATGGCGAATGGGACGCGCCCTGTAGCGGC
GCATTAAGCGCGGCGGGTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCC
CTAGCGCCCGCTCCTTTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCC
CGTCAAGCTCTAAATCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTC
GACCCCAAAAAACTTGATTAGGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACG
GTTTTTCGCCCTTTGACGTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACT
GGAACAACACTCAACCCTATCTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATT
TCGGCCTATTGGTTAAAAAATGAGCTGATTTAACAAAAATTTAACGCGAATTTTAACAAA
ATATTAACGCTTACAATTTAGGTGGCACTTTTCGGGGAAATGTGCGCGGAACCCCTATTT
GTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGACAATAACCCTGATAAA
TGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCCGTGTCGCCCTTA
TTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAG
TAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACA
GCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCACTTTTA
AAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTC
GCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATC
TTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACA
CTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGC
ACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCA
TACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAAC
TATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGG
CGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTG
ATAAATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATG
GTAAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAAC
GAAATAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACC
AAGTTTACTCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCT
AGGTGAAGATCCTTTTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCC
ACTGAGCGTCAGACCCCGTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGC
GCGTAATCTGCTGCTTGCAAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGG
ATCAAGAGCTACCAACTCTTTTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAA
ATACTGTCCTTCTAGTGTAGCCGTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGC
CTACATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGT
GTCTTACCGGGTTGGACTCAAGACGATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAA
CGGGGGGTTCGTGCACACAGCCCAGCTTGGAGCGAACGACCTACACCGAACTGAGATACC
TACAGCGTGAGCTATGAGAAAGCGCCACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATC
CGGTAAGCGGCAGGGTCGGAACAGGAGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCT
GGTATCTTTATAGTCCTGTCGGGTTTCGCCACCTCTGACTTGAGCGTCGATTTTTGTGAT
GCTCGTCAGGGGGGCGGAGCCTATGGAAAAACGCCAGCAACGCGGCCTTTTTACGGTTCC
TGGCCTTTTGCTGGCCTTTTGCTCACATGTTCTTTCCTGCGTTATCCCCTGATTCTGTGG
ATAACCGTATTACCGCCTTTGAGTGAGCTGATACCGCTCGCCGCAGCCGAACGACCGAGC
GCAGCGAGTCAGTGAGCGAGGAAGCGGAAGAGCGCCCAATACGCAAACCGCCTCTCCCCG
CGCGTTGGCCGATTCATTAATGCAGCTGGCACGACAGGTTTCCCGACTGGAAAGCGGGCA
GTGAGCGCAACGCAATTAATGTGAGTTAGCTCACTCATTAGGCACCCCAGGCTTTACACT
TTATGCTTCCGGCTCGTATGTTGTGTGGAATTGTGAGCGGATAACAATTTCACACAGGAA
ACAGCTATGACCATGATTACGCCAAGCTCGAAATTAACCCTCACTAAAGGGAACAAAAGC
TGGTACCGTTGCCATTAGTATGAAGAAATAGTAAAAATGTTTTACACATTTAGAGGCATT
CCCTATATGGATAACTTCGATTTTACGGGTATATGAAAAATAATCAATTAAGCATTTTAA
TATATTATTATTATTATTATTATTATTATTATTACTCTCATGTTATATTCATTGTTTTAT
TTTTTAATTTTATATGCTACCATTTTTCATCCGTCATATGTCATTTTTTGTAATATATTA
ACTTACATGATGAAAAATTATAATTTTTTTTAGAAAAATTGGTTCACGAATATTATTCAA
ATTTATCATTGATGTATAATAGTTATGAAAGCCGAACCACACCATTCTGTGGATATAGTT
TTGTTTTATCTGCAGATATTCAAGTAAGGAAAACTTCTTTTCATTTACCCCTCTATATAT
CATATTTTTCATATTTACACAATTTATTCAAATTCAAATACATTTGTATTAATTTTGTAG
GTATCAGAAGACATAACAATGTTTGTTCAAACATCAGTAGATATAGATGACCACAACAAG
CTTGGGCCCCCGCGGTGGCGGCCGCTCTAGCTTTGATCCCGTTTTTCTTACTTATATATT
TATACCAATTGATTGTATTTATAACTGTAAAAATGTGTATGTTGTGTGCATATTTTTTTT
TGTGCATGCACATGCATGTAAATAGCTAAAATTATGAACATTTTATTTTTTGTTCAGAAA
AAAAAAACTTTACACACATAAAATGGCTAGTATGAATAGCCATATTTTATATAAATTAAA
TCCTATGAATTTATGACCATATTAAAAATTTAGATATTTATGGAACATAATATGTTTGAA
ACAATAAGACAAAATTATTATTATTATTATTATTTTTACTGTTATAATTATGTTGTCTCT
TCAATGATTCATAAATAGTTGGACTTGATTTTTAAAATGTTTATAATATGATTAGCATAG
TTAAATAAAAAAAGTTGAAAAATTAAAAAAAAACATATAAACACAAATGATGTTTTTTCC
TTCAATTTCGATTGATAATTCCTGCAGCCCAGCTTAATTCTTTTCGAGCTCTTTATGCTT
AAGTTTACAATTTAATATTCATACTTTAAGTATTTTTTGTAGTATCCTAGATATTGTGCT
TTAAATGCTCACCCCTCAAAGCACCAGTAATATTTTCATCCACTGAAATACCATTAAATT
TTCAAAAAAATACTATGCATATAATGTTATACATATAAACATAAAACGCCATGTAAATCA
AAAAATATATAAAAATATGTATAAAAATAAATATGCACTAAATATAAGCTAATTATGCAT
AAAAATTAAAGTGCCCTTTATTAACTAGAACTAGTCGTAATTATTTATATTTCTATGTTA
TAAAAAAATCCTCATATAATAATATAATTAATATATGTAATGTTTTTTTTATTTTATAAT
TTTAATATAAAATAATATGTAAATTAATTCAAAAAATAAATATAATTGTTGTGAAACAAA
AAACGTAATTTTTTCATTTGCCTTCAAAATTTAAATTTATTTTAATATTTCCTAAAATAT
ATATACTTTGTGTATAAATATATAAAAATATATATTTGCTTATAAATAAATAAAAAATTT
TATAAAACATAGGGGGATCCATGGTTGGTTCGCTAAACTGCATCGTCGCTGTGTCCCAGA
ACATGGGCATCGGCAAGAACGGGGACCTGCCCTGGCCACCGCTCAGGAACGAATTTAGAT
ATTTCCAGAGAATGACCACAACCTCTTCAGTAGAAGGTAAACAGAATCTGGTGATTATGG
GTAAGAAGACCTGGTTCTCCATTCCTGAGAAGAATCGACCTTTAAAGGGTAGAATTAATT
TAGTTCTCAGCAGAGAACTCAAGGAACCTCCACAAGGAGCTCATTTTCTTTCCAGAAGTC
TAGATGATGCCTTAAAACTTACTGAACAACCAGAATTAGCAAATAAAGTAGACATGGTCT
GGATAGTTGGTGGCAGTTCTGTTTATAAGGAAGCCATGAATCACCCAGGCCATCTTAAAC
TATTTGTGACAAGGATCATGCAAGACTTTGAAAGTGACACGTTTTTTCCAGAAATTGATT
TGGAGAAATATAAACTTCTGCCAGAATACCCAGGTGTTCTCTCTGATGTCCAGGAGGAGA
AAGGCATTAAGTACAAATTTGAAGTATATGAGAAGAATGATGCTAGCGGAGGAGGTGGAT
CTGGTGGAGGTGGAAGTGCTAGCGTGACAGGGGGAATGGCAAGCAAGTGGGATCAGAAGG
GTATGGACATTGCCTATGAGGAGGCGGCCTTAGGTTACAAAGAGGGTGGTGTTCCTATTG
GCGGATGTCTTATCAATAACAAAGACGGAAGTGTTCTCGGTCGTGGTCACAACATGAGAT
TTCAAAAGGGATCCGCCACACTACATGGTGAGATCTCCACTTTGGAAAACTGTGGGAGAT
TAGAGGGCAAAGTGTACAAAGATACCACTTTGTATACGACGCTGTCTCCATGCGACATGT
GTACAGGTGCCATCATCATGTATGGTATTCCACGCTGTGTTGTCGGTGAGAACGTTAATT
TCAAAAGTAAGGGCGAGAAATATTTACAAACTAGAGGTCACGAGGTTGTTGTTGTTGACG
ATGAGAGGTGTAAAAAGATCATGAAACAATTTATCGATGAAAGACCTCAGGATTGGTTTG
AAGATATTGGTGAGGCTTCGGAACCATTTAAGAACGTCTACTTGCTACCTCAAACAAACC
AATTGCTGGGTTTGTACACCATCATCAGAAATAAGAATACAACTAGACCTGATTTCATTT
TCTACTCCGATAGAATCATCAGATTGTTGGTTGAAGAAGGTTTGAACCATCTACCTGTGC
AAAAGCAAATTGTGGAAACTGACACCAACGAAAACTTCGAAGGTGTCTCATTCATGGGTA
AAATCTGTGGTGTTTCCATTGTCAGAGCTGGTGAATCGATGGAGCAAGGATTAAGAGACT
GTTGTAGGTCTGTGCGTATCGGTAAAATTTTAATTCAAAGGGACGAGGAGACTGCTTTAC
CAAAGTTATTCTACGAAAAATTACCAGAGGATATATCTGAAAGGTATGTCTTCCTATTAG
ACCCAATGCTGGCCACCGGTGGTAGTGCTATCATGGCTACAGAAGTCTTGATTAAGAGAG
GTGTTAAGCCAGAGAGAATTTACTTCTTAAACCTAATCTGTAGTAAGGAAGGGATTGAAA
AATACCATGCCGCCTTCCCAGAGGTCAGAATTGTTACTGGTGCCCTCGACAGAGGTCTAG
ATGAAAACAAGTATCTAGTTCCAGGGTTGGGTGACTTTGGTGACAGATACTACTGTGTTT
AACTCGATCCCGTTTTTCTTACTTATATATTTATACCAATTGATTGTATTTATAACTGTA
AAAATGTGTATGTTGTGTGCATATTTTTTTTTGTGCATGCACATGCATGTAAATAGCTAA
AATTATGAACATTTTATTTTTTGTTCAGAAAAAAAAAACTTTACACACATAAAATGGCTA
GTATGAATAGCCATATTTTATATAAATTAAATCCTATGAATTTATGACCATATTAAAAAT
TTAGATATTTATGGAACATAATATGTTTGAAACAATAAGACAAAATTATTATTATTATTA
TTATTTTTACTGTTATAATTATGTTGTCTCTTCAATGATTCATAAATAGTTGGACTTGAT
TTTTAAAATGTTTATAATATGATTAGCATAGTTAAATAAAAAAAGTTGAAAAATTAAAAA
AAAACATATAAACACAAATGATGTTTTTTCCTTCAATTTCGGGTACGTACCCTCGAGGCT
AGCGAT
Restriction sites to linearize plasmid Asp718I/EcoRI
Partial or complete disruption of the geneUnknown
Additional remarks partial/complete disruption
Selectable marker used to select the mutant parasitehdhfr
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 1GCCGGTACCGTTGCCATTAGTATGAAGAAATAG
Additional information primer 1F1 primer
Sequence Primer 2GCCAAGCTTGTTGTGGTCATCTATATCTACTGATG
Additional information primer 2R1 primer
Sequence Primer 3GCCGATATCGCGACAGTGTTCGTGTTACATACG
Additional information primer 3F2 primer
Sequence Primer 4GCCGATATCGCGACAGTGTTCGTGTTACATACG
Additional information primer 4R2 primer
Sequence Primer 5
Additional information primer 5
Sequence Primer 6
Additional information primer 6
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Conceptual design of the database: Dr. C.J. Janse (Leiden Malaria Research Group, Leiden, The Netherlands).
Technical design and realization: S. Mededovic and A. van Wigcheren