RMgmDB - Rodent Malaria genetically modified Parasites

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Summary

RMgm-4057
Malaria parasiteP. yoelii
Genotype
DisruptedGene model (rodent): PY17X_1030600; Gene model (P.falciparum): PF3D7_1414500; Gene product: atypical protein kinase, ABC-1 family, putative (ABCk2)
Transgene
 Transgene not Plasmodium: GFP-Luciferase
Promoter: Gene model: PBANKA_1133300; Gene model (P.falciparum): PF3D7_1357100; Gene product: elongation factor 1-alpha (eef1a)
3'UTR: Gene model: PBANKA_0719300; Gene product: bifunctional dihydrofolate reductase-thymidylate synthase, putative (dhfr/ts)
Replacement locus: Gene model: PY17X_0306600; Gene product: 6-cysteine protein (P230p)
PhenotypeNo phenotype has been described
Last modified: 24 December 2016, 17:35
  *RMgm-4057
Successful modificationThe parasite was generated by the genetic modification
The mutant contains the following genetic modification(s) Gene disruption, Introduction of a transgene
Reference (PubMed-PMID number) Not published (yet)
MR4 number
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Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. yoelii
Parent strain/lineP. y. yoelii 17XNL
Name parent line/clone RMgm-689
Other information parent lineA reference transgenic line of P. y. yoelii 17XNL (1971cl1) expressing GFP-Luciferase under control of the constitutive P. berghei eef1a promoter. The gfp-luciferase expression cassette is introduced into the silent 230p locus.
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The mutant parasite was generated by
Name PI/ResearcherFranke-Fayard B; Janse CJ
Name Group/DepartmentLeiden Malaria Research Group
Name InstituteLeiden University Medical Center
CityLeiden
CountryThe Netherlands
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Name of the mutant parasite
RMgm numberRMgm-4057
Principal name2753cl3
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 different from wild type
Fertilization and ookineteNot tested
OocystNot different from wild type
SporozoiteNot different from wild type
Liver stageNot different from wild type
Additional remarks phenotype

Mutant/mutation
The mutant lacks expression of atypical protein kinase, ABC-1 family, putative and expresses GFP-Luciferase under control of the constitutive P. berghei eef1a promoter

Protein (function)

Phenotype
The phenotype of blood stages (growth rate), oocyst and sporozoite production and liver stage development were not different from wild type parasites. Liver stage development was analysed by measuring prepatent period after infection of mice with defined numbers of sporozoites either by intravenous or intradermal administration. 

Additional information

Below data of genotype analysis of parasites of line 2753



Figure: Genotype analysis of line 2753 by Southern analysis of pulsed field gel separated chromosomes (upper panel) and diagnostic PCR (lower panel) confirming correct integration of the deletion construct that integrates by double cross-over integration into the abck2 locus. Separated chromosomes are hybridized with a probe recognizing the 3'-UTR of the pbdhfr/ts gene recognizing the endogenous dhfr/ts gene on chromosome 7 and the 3'-UTR dhfr/ts of the integrated construct at chromosome 10 (right panel: parasites of line 2753 uncloned; right panel parasites of clone 3 of line 2753). SM: selectable marker; ORF: Open reading frame; 3' and 5': 3'and 5'-integration of the construct.

Other mutants

  Disrupted: Mutant parasite with a disrupted gene
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Details of the target gene
Gene Model of Rodent Parasite PY17X_1030600
Gene Model P. falciparum ortholog PF3D7_1414500
Gene productatypical protein kinase, ABC-1 family, putative
Gene product: Alternative nameABCk2
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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
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Plasmid/construct sequence
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AATTCACTGGCCGTCGTTTTACAACGTCGTGACTGGGAAAACCCTGGCGTTACCCAACTT
AATCGCCTTGCAGCACATCCCCCTTTCGCCAGCTGGCGTAATAGCGAAGAGGCCCGCACC
GATCGCCCTTCCCAACAGTTGCGCAGCCTGAATGGCGAATGGCGCCTGATGCGGTATTTT
CTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGC
TCTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGA
CGGGCTTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGC
ATGTGTCAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATA
CGCCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACT
TTTCGGGGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATG
TATCCGCTCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGT
ATGAGTATTCAACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCT
GTTTTTGCTCACCCAGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCA
CGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCC
GAAGAACGTTTTCCAATGATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCC
CGTATTGACGCCGGGCAAGAGCAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTG
GTTGAGTACTCACCAGTCACAGAAAAGCATCTTACGGATGGCATGACAGTAAGAGAATTA
TGCAGTGCTGCCATAACCATGAGTGATAACACTGCGGCCAACTTACTTCTGACAACGATC
GGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACATGGGGGATCATGTAACTCGCCTT
GATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGACGAGCGTGACACCACGATG
CCTGTAGCAATGGCAACAACGTTGCGCAAACTATTAACTGGCGAACTACTTACTCTAGCT
TCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGTTGCAGGACCACTTCTGCGC
TCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCGGTGAGCGTGGGTCT
CGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCCTCCCGTATCGTAGTTATCTAC
ACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACAGATCGCTGAGATAGGTGCC
TCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATATATACTTTAGATTGAT
TTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGATAATCTCATG
ACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAGAAAAGATC
AAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAAA
CCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCGAAG
GTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTCCTTCTAGTGTAGCCGTAGTTA
GGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTA
CCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAG
TTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTG
GAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCATTGAGAAAGCGCCACG
CTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAG
CGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGC
CACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAA
AACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATG
TTCTTTCCTGCGTTATCCCCTGATTCTGTGGATAACCGTATTACCGCCTTTGAGTGAGCT
GATACCGCTCGCCGCAGCCGAACGACCGAGCGCAGCGAGTCAGTGAGCGAGGAAGCGGAA
GAGCGCCCAATACGCAAACCGCCTCTCCCCGCGCGTTGGCCGATTCATTAATGCAGCTGG
CACGACAGGTTTCCCGACTGGAAAGCGGGCAGTGAGCGCAACGCAATTAATGTGAGTTAG
CTCACTCATTAGGCACCCCAGGCTTTACACTTTATGCTTCCGGCTCGTATGTTGTGTGGA
ATTGTGAGCGGTAAACAATTTCACACAGGAAACAGCTTGACCATGATTACGCCAAGCTTG
CCATTTTGTCATTTTGCCATTTTGCCATTTTGCCATTTTTTTGTTTTATTTTGCCATTTT
TTTATTTTGTTTTGTCTTATTCCAATGTGCTAATACATTGCACTATTATTTTATGTTTAT
AATTGTTACTAACAAATGTATGATAAAAAAGAAGATATGAATGGTTTATTTGTTTCGGTA
TAATTTGTATATTTATTAATTTTCTTTTATTTTTTTGTATTAATTTTTGGAATTTTGTCT
AGACATTTGTATTTTAGTAGCAGCCTATTAATATCCTTATACAAACATTTTTATGTGTTT
TTATAATTGTAGATATTTTATTTATTTGTTTTGTGTAAAATGATTTTTAATATATGCACA
TGCATATAGAGAATAATATCATTTGTACATTTTTCGTTAACATATGTATGTATGTATATA
TTTTTTTTTTTTAATTACATGTTGTTAATCTTTGTTTAGTTCTTGTTTTTACATTTTTAT
GTTATTTTTTACTTTGTTTTTGCCAATTATAAAAAATACATAGAAATGTTAATATATTAT
AGAATAACCACTAAATTCATTTCTTTTTAAATAATTATACTATCATAATATATACATTTG
AATAATACTACATTTGAATAATACTACATTGTTAGATTATTTGATATCATTTTTAATAAA
TGAAAATGGTTAGCATTTTTTTTTTTTACTAGCTATTTTGGTATTTCCTTGCAGGCCTCT
GCAGCCCAGCTTAATTCTTTTCGAGCTCTTTATGCTTAAGTTTACAATTTAATATTCATA
CTTTAAGTATTTTTTGTAGTATCCTAGATATTGTGCTTTAAATGCTCACCCCTCAAAGCA
CCAGTAATATTTTCATCCACTGAAATACCATTAAATTTTCAAAAAAATACTATGCATATA
ATGTTATACATATAAACATAAAACGCCATGTAAATCAAAAAATATATAAAAATATGTATA
AAAATAAATATGCACTAAATATAAGCTAATTATGCATAAAAATTAAAGTGCCCTTTATTA
ACTAGTCGTAATTATTTATATTTCTATGTTATAAAAAAATCCTCATATAATAATATAATT
AATATATGTAATGTTTTTTTTATTTTATAATTTTAATATAAAATAATATGTAAATTAATT
CAAAAAATAAATATAATTGTTGTGAAACAAAAAACGTAATTTTTTCATTTGCCTTCAAAA
TTTAAATTTATTTTAATATTTCCTAAAATATATATACTTTGTGTATAAATATATAAAAAT
ATATATTTGCTTATAAATAAATAAAAAATTTTATAAAACATAGGGGGATCCATGGTTGGT
TCGCTAAACTGCATCGTCGCTGTGTCCCAGAACATGGGCATCGGCAAGAACGGGGACCTG
CCCTGGCCACCGCTCAGGAACGAATTTAGATATTTCCAGAGAATGACCACAACCTCTTCA
GTAGAAGGTAAACAGAATCTGGTGATTATGGGTAAGAAGACCTGGTTCTCCATTCCTGAG
AAGAATCGACCTTTAAAGGGTAGAATTAATTTAGTTCTCAGCAGAGAACTCAAGGAACCT
CCACAAGGAGCTCATTTTCTTTCCAGAAGTCTAGATGATGCCTTAAAACTTACTGAACAA
CCAGAATTAGCAAATAAAGTAGACATGGTCTGGATAGTTGGTGGCAGTTCTGTTTATAAG
GAAGCCATGAATCACCCAGGCCATCTTAAACTATTTGTGACAAGGATCATGCAAGACTTT
GAAAGTGACACGTTTTTTCCAGAAATTGATTTGGAGAAATATAAACTTCTGCCAGAATAC
CCAGGTGTTCTCTCTGATGTCCAGGAGGAGAAAGGCATTAAGTACAAATTTGAAGTATAT
GAGAAGAATGATGCTAGCGGAGGAGGTGGATCTGGTGGAGGTGGAAGTGCTAGCGTGACA
GGGGGAATGGCAAGCAAGTGGGATCAGAAGGGTATGGACATTGCCTATGAGGAGGCGGCC
TTAGGTTACAAAGAGGGTGGTGTTCCTATTGGCGGATGTCTTATCAATAACAAAGACGGA
AGTGTTCTCGGTCGTGGTCACAACATGAGATTTCAAAAGGGATCCGCCACACTACATGGT
GAGATCTCCACTTTGGAAAACTGTGGGAGATTAGAGGGCAAAGTGTACAAAGATACCACT
TTGTATACGACGCTGTCTCCATGCGACATGTGTACAGGTGCCATCATCATGTATGGTATT
CCACGCTGTGTTGTCGGTGAGAACGTTAATTTCAAAAGTAAGGGCGAGAAATATTTACAA
ACTAGAGGTCACGAGGTTGTTGTTGTTGACGATGAGAGGTGTAAAAAGATCATGAAACAA
TTTATCGATGAAAGACCTCAGGATTGGTTTGAAGATATTGGTGAGGCTTCGGAACCATTT
AAGAACGTCTACTTGCTACCTCAAACAAACCAATTGCTGGGTTTGTACACCATCATCAGA
AATAAGAATACAACTAGACCTGATTTCATTTTCTACTCCGATAGAATCATCAGATTGTTG
GTTGAAGAAGGTTTGAACCATCTACCTGTGCAAAAGCAAATTGTGGAAACTGACACCAAC
GAAAACTTCGAAGGTGTCTCATTCATGGGTAAAATCTGTGGTGTTTCCATTGTCAGAGCT
GGTGAATCGATGGAGCAAGGATTAAGAGACTGTTGTAGGTCTGTGCGTATCGGTAAAATT
TTAATTCAAAGGGACGAGGAGACTGCTTTACCAAAGTTATTCTACGAAAAATTACCAGAG
GATATATCTGAAAGGTATGTCTTCCTATTAGACCCAATGCTGGCCACCGGTGGTAGTGCT
ATCATGGCTACAGAAGTCTTGATTAAGAGAGGTGTTAAGCCAGAGAGAATTTACTTCTTA
AACCTAATCTGTAGTAAGGAAGGGATTGAAAAATACCATGCCGCCTTCCCAGAGGTCAGA
ATTGTTACTGGTGCCCTCGACAGAGGTCTAGATGAAAACAAGTATCTAGTTCCAGGGTTG
GGTGACTTTGGTGACAGATACTACTGTGTTTAACTCGATCCCGTTTTTCTTACTTATATA
TTTATACCAATTGATTGTATTTATAACTGTAAAAATGTGTATGTTGTGTGCATATTTTTT
TTTGTGCATGCACATGCATGTAAATAGCTAAAATTATGAACATTTTATTTTTTGTTCAGA
AAAAAAAAACTTTACACACATAAAATGGCTAGTATGAATAGCCATATTTTATATAAATTA
AATCCTATGAATTTATGACCATATTAAAAATTTAGATATTTATGGAACATAATATGTTTG
AAACAATAAGACAAAATTATTATTATTATTATTATTTTTACTGTTATAATTATGTTGTCT
CTTCAATGATTCATAAATAGTTGGACTTGATTTTTAAAATGTTTATAATATGATTAGCAT
AGTTAAATAAAAAAAGTTGAAAAATTAAAAAAAAACATATAAACACAAATGATGTTTTTT
CCTTCAATTTCGGGTACCCTCGAGCAGACAAATTTGATAAATCAGACAAATTTGATAAGT
CAGACAAATTTGATAAGTCAGACAAATTTGATAAGTCAGACAAATTTGATAAGTCAGACA
AATTTGATAAGTCAGACAAATTTGATAAATTGCAAAGTTTTAAAGATTCAGAAAAAAAAA
AAAATAATAAAGAAGAGAACTTTTATGATGCAAAAGATGATATAACAGAAGATAATATAA
ATAACAGATGTAATGACAAAATAAAAAATAATATTAAGCAAAAAGATATTTTTAGTCTTA
AACATACATTCCTTAATCATGTTGATAATTTAAAAGAAAAGACAAAAAAAATTAATGATT
CGATAAAAAATATATATGAAAATAATGATAACATTTTTTTAAAGAATTTATTTACATCTC
AAAAAAAAGCAGAAAAAGGGGAAAAATATAATAATAGCCATTTTGATAATAGTACCCATT
TTGATAGTAGTGCCAATTTCGATAATAATAACGATTTAGAAAAAGATTATGATATAAGAA
ATATAGAATATAAACAGTTATATGAAAAAAAAAAATTAAAAAAAAATAAAAAAAGTGATA
TTTTTTTTGAAAATATTTTAAAAAATGGAAAAATTAATAATGTTGACAAAATTAATAATG
TTGACAAAATTTTGAACATAAATAAACTAAAATCACCACTTAAGATATTAGAAAAAAATA
AAAAAAAAATAAATATAAGAAATAATTCCAATAGAAGATCATTAAGTTATAATAATATTG
ATCATGAAAATAATTTATTTAAAAATTTAGAATCAATTAATGATACAAATCATGATTGTA
ATCAAAATATGCCAAAGTTTACAAATGAAAATGATGAAAATGATGAAAATGATGAAAATA
AAAAAGGAAAATTAGATTATAATAAAAATTCACAAAATGAATTGATTCCATCCATTTTTA
TAAACAACATTATTTTAGAATCTAAAGTAGACAAGGGTGGAAATATAGG
Restriction sites to linearize plasmid EcoRI, HindIII, PvuI
Partial or complete disruption of the geneComplete
Additional remarks partial/complete disruption
Selectable marker used to select the mutant parasitehdhfr/yfcu
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
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  Transgene: Mutant parasite expressing a transgene
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Type and details of transgene
Is the transgene Plasmodium derived Transgene: not Plasmodium
Transgene nameGFP-Luciferase
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Details of the genetic modification
Inducable system usedNo
Additional remarks inducable system
Type of plasmid/construct(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
Selectable marker used to select the mutant parasitehdhfr/yfcu
Promoter of the selectable markereef1a
Selection (positive) procedureNo
Selection (negative) procedure5-fluorocytosine (5-FC)
Additional remarks genetic modification
Additional remarks selection procedure
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Other details transgene
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Promoter
Gene Model of Parasite PBANKA_1133300
Gene Model P. falciparum ortholog PF3D7_1357100
Gene productelongation factor 1-alpha
Gene product: Alternative nameeef1a
Primer information details of the primers used for amplification of the promoter sequence  Click to view information
Primer information details of the primers used for amplification of the promoter sequence  Click to hide information
Sequence Primer 1
Additional information primer 1
Sequence Primer 2
Additional information primer 2
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3'-UTR
Gene Model of Parasite PBANKA_0719300
Gene productbifunctional dihydrofolate reductase-thymidylate synthase, putative
Gene product: Alternative namedhfr/ts
Primer information details of the primers used for amplification the 3'-UTR sequences  Click to view information
Primer information details of the primers used for amplification the 3'-UTR sequences  Click to hide information
Sequence Primer 1
Additional information primer 1
Sequence Primer 2
Additional information primer 2
Insertion/Replacement locus
Replacement / InsertionReplacement locus
Gene Model of Parasite PY17X_0306600
Gene product6-cysteine protein
Gene product: Alternative nameP230p
Primer information details of the primers used for amplification of the target sequences  Click to view information
Primer information details of the 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
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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