Summary

RMgm-707
Malaria parasiteP. berghei
Genotype
DisruptedGene model (rodent): PBANKA_0100600; Gene model (P.falciparum): Not available; Gene product: schizont membrane associated cytoadherence protein (Schizont Membrane Associated Cytoadherence protein; SMAC)
TaggedGene model (rodent): PBANKA_1365500; Gene model (P.falciparum): Not available; Gene product: intra-erythrocytic P. berghei-induced structures protein 1 exported protein IBIS1 (Plasmodium exported protein IBIS1)
Name tag: mCherry
Transgene
Transgene not Plasmodium: A fusion of GFP (gfp-mu3) and Luciferase Firefly (LucIAV)
Promoter: Gene model: PBANKA_0915000; Gene model (P.falciparum): PF3D7_1133400; Gene product: apical membrane antigen 1 (AMA-1)
3'UTR: Gene model: PBANKA_0719300; Gene product: bifunctional dihydrofolate reductase-thymidylate synthase, putative (dhfr/ts)
Replacement locus: Gene model: PBANKA_0306000; Gene product: 6-cysteine protein (230p)
Phenotype Asexual bloodstage;
Last modified: 2 December 2012, 15:33
  *RMgm-707
Successful modificationThe parasite was generated by the genetic modification
The mutant contains the following genetic modification(s) Gene disruption, Gene tagging, Introduction of a transgene
Reference (PubMed-PMID number) Reference 1 (PMID number) : 23197789
MR4 number
Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. berghei
Parent strain/lineP. berghei ANKA
Name parent line/clone 1242cl5m2cl1
Other information parent line1242cl5m2cl1 (RMgm-662) lacks expression of Schizont Membrane Associated Cytoadherence protein (SMAC) and expresses GFP-luciferase under control of a schizont-specific promoter. This line does not contain a drug selectable marker (PubMed: PMID: 22184632)
The mutant parasite was generated by
Name PI/ResearcherE.M. Pasini, C.J. Janse, B.M.D. Franke-Fayard
Name Group/DepartmentDepartment of Parasitology
Name InstituteBiomedical Primate Research Centre
CityRijswijk
CountryThe Netherlands
Name of the mutant parasite
RMgm numberRMgm-707
Principal name2071; 2072
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Phenotype
Asexual blood stageThe tagged-protein is exported into the cytoplasm of the host erythrocyte and shows a punctuate, vesicle-like localization in the cytoplasm
Gametocyte/GameteNot tested
Fertilization and ookineteNot tested
OocystNot tested
SporozoiteNot tested
Liver stageNot tested
Additional remarks phenotype

Mutant/mutation

Protein (function)

Phenotype

Additional information
The mutant parasite expresses an mCherry-tagged protein. The protein has been selected for tagging in a screen for putative exported proteins of P. berghei.

The genotype of the parasites has not been analyzed in detail.
Southern analysis of PFG-separated chromosomes (to show integration into the chromosome on which the target gene is located) has been performed (see below, no details available for mutant 2009cl1). This analysis provided evidence for tagging of the correct gene. The construct used aims at integration of the tagging construct by single-cross-over integration resulting in the presence of a tagged copy of the endogenous gene.

Phenotype analyses of the blood stages by fluorescence microscopy show that the tagged-protein is exported into the cytoplasm of the host erythrocyte and shows a punctuate, vesicle-like localization in the cytoplasm.

Additional mutants were made in P. berghei ANKA cl15cy1 expressing either an mCherry-tagged (1951), or EGFP-tagged (2009cl1) version of PBANKA_136650. P. berghei ANKA cl15cy1 is a reference wild type clone from the ANKA strain of P. berghei (PubMed: PMID: 17406255).



Other mutants
RMgm-705: mutant expressing GFP-luciferase under control of a schizont-specific promoter and expressing an mCherry-tagged version of PBANKA_136550
RMgm-706: P. berghei K173 mutant expressing an mCherry-tagged version of PBANKA_136550
RMgm-717: mutant lacking expression of PBANKA_136550


  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_0100600
Gene Model P. falciparum ortholog Not available
Gene productschizont membrane associated cytoadherence protein
Gene product: Alternative nameSchizont Membrane Associated Cytoadherence protein; SMAC
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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ATCGAATTCCTAAGATCATTATGGGATCATCTGCAGCGATATTTTTTCTTGTCGAAGATT
TAGGAATTAGAGCTGTGCTATTTTTAATATTTTCTGCTGCTGTACTCGCTTATTTCACTT
CCATATAAATTAAAAGAACAAACACATATTTATATATTAAAATAATAATAATATTCTTTT
ACTATTATTAATGAGATAAATATGAAATTATGCATATTTAACTAATAAAAAATACACAAT
GTGCATTATAAATATAGCAATCGTTACAATATATTTACATTATTAATATATTTTCATTAT
TTGTGAATGTTAATAATTAAGTGTTATGGATATGGTAGATTATTTTATACTTGAGACAGT
GTGCTTATGCATCTATGTATTAATTGTTATTATTAAAATTTATAATAAAGTGCAATAAGC
TATATTTTGAAATATTTCCTATATAAATATATACATTTTATGAGTTTATAATTCTATATA
CATTCTTGTATTAATATTAAATGAATTTTTTTTTGTGAAAAAACATAAATAATATATAGT
GATAATACACAAATGATTGAAACGTGAGGTGAAACATCTTTTCATTTATGAATACAACTC
TTTAAGTATGTTTTTTTTTCAGTGAAAACGCAAACTTGTATAGAATATATTTTTTTTATT
AGTTCTCCATCATTATTTATATAGTTTTATCTCCCCATAAAATATTAATAATACAACCCT
CTAGAGCGGCCGCCACCGCGGTGGAGCTCCAATTCGCCCTATAGTGAGTCGTATTACAAT
TCACTGGCCGTCGTTTTACAACGTCGTGACTGGGAAAACCCTGGCGTTACCCAACTTAAT
CGCCTTGCAGCACATCCCCCTTTCGCCAGCTGGCGTAATAGCGAAGAGGCCCGCACCGAT
CGCCCTTCCCAACAGTTGCGCAGCCTGAATGGCGAATGGGACGCGCCCTGTAGCGGCGCA
TTAAGCGCGGCGGGTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTA
GCGCCCGCTCCTTTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGT
CAAGCTCTAAATCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGAC
CCCAAAAAACTTGATTAGGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTT
TTTCGCCCTTTGACGTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGA
ACAACACTCAACCCTATCTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCG
GCCTATTGGTTAAAAAATGAGCTGATTTAACAAAAATTTAACGCGAATTTTAACAAAATA
TTAACGCTTACAATTTAGGTGGCACTTTTCGGGGAAATGTGCGCGGAACCCCTATTTGTT
TATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGACAATAACCCTGATAAATGC
TTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCCGTGTCGCCCTTATTC
CCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAGTAA
AAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACAGCG
GTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCACTTTTAAAG
TTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTCGCC
GCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTA
CGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACACTG
CGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACA
ACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATAC
CAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTAT
TAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGG
ATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATA
AATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTA
AGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAA
ATAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAG
TTTACTCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGG
TGAAGATCCTTTTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACT
GAGCGTCAGACCCCGTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCG
TAATCTGCTGCTTGCAAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATC
AAGAGCTACCAACTCTTTTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATA
CTGTCCTTCTAGTGTAGCCGTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTA
CATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTC
TTACCGGGTTGGACTCAAGACGATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGG
GGGGTTCGTGCACACAGCCCAGCTTGGAGCGAACGACCTACACCGAACTGAGATACCTAC
AGCGTGAGCTATGAGAAAGCGCCACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGG
TAAGCGGCAGGGTCGGAACAGGAGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGT
ATCTTTATAGTCCTGTCGGGTTTCGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCT
CGTCAGGGGGGCGGAGCCTATGGAAAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGG
CCTTTTGCTGGCCTTTTGCTCACATGTTCTTTCCTGCGTTATCCCCTGATTCTGTGGATA
ACCGTATTACCGCCTTTGAGTGAGCTGATACCGCTCGCCGCAGCCGAACGACCGAGCGCA
GCGAGTCAGTGAGCGAGGAAGCGGAAGAGCGCCCAATACGCAAACCGCCTCTCCCCGCGC
GTTGGCCGATTCATTAATGCAGCTGGCACGACAGGTTTCCCGACTGGAAAGCGGGCAGTG
AGCGCAACGCAATTAATGTGAGTTAGCTCACTCATTAGGCACCCCAGGCTTTACACTTTA
TGCTTCCGGCTCGTATGTTGTGTGGAATTGTGAGCGGATAACAATTTCACACAGGAAACA
GCTATGACCATGATTACGCCAAGCTCGAAATTAACCCTCACTAAAGGGAACAAAAGCTGG
TACCGTGTATATATAATCAAATACCAAGGATAATATCTATATATTTATATTTTCTAAAAG
AGACATTTTAGATATAATATAAACTTTATTGTAATTATTACATTTCACGTAAAAATATAT
AATTTTTTTCGAGGAAGTTTTAAATAAAATATTTATATGATATATTTATTAATTTTTTAT
TTAATTAGTAATTTGTTACATTTATATATTGAACCCATTTTAAAAATATACATATTAAAT
AGCTTTAGAATTATGACAATTAATTTTTATAATAATAAAAAGTTCCATTTATTTGTTTTC
TGAATCGGTAAAGAGTATTAATAAAAACCCAATTGTAAACGGTCCCAATATATTTTTTAA
AGAATGTGTTACTATGTATTTAAATTACCCAAATTAAACAAGTAAAAAATATCACAATAT
ATATTATTTTCCCCGAGATGCTATAAATAATTATTTTTGATAGATTGACAAATATATATT
GTTCTTTTTTGACGTAATGCATTATATATCATATATGTAGCACTTTTTAGTTTTAAAATA
ATAATTTAATATAATACACAGTGCATGCACCATTTTACATGAATCTGATATAATACTTAT
TTATCATTTTATTTATTAGCCATTTATTATAAACAAACTATATATTATTATCTTTTACAT
ACATCCTTAACTCTTTTAACAACCAGTACACATAATATTTGCAACATTGTGAATATAAAA
TGAAATATTTAACGCACATATTTTTTTTTATTATACTTTTTCAATTTATAGTTGATAAAT
ACGATAACAATGGAAAATCATTGGACAAGAAGCTTGGGCCCCCGCGGTGGCGGCCGCTCT
AGCTTTGATCCCGTTTTTCTTACTTATATATTTATACCAATTGATTGTATTTATAACTGT
AAAAATGTGTATGTTGTGTGCATATTTTTTTTTGTGCATGCACATGCATGTAAATAGCTA
AAATTATGAACATTTTATTTTTTGTTCAGAAAAAAAAAACTTTACACACATAAAATGGCT
AGTATGAATAGCCATATTTTATATAAATTAAATCCTATGAATTTATGACCATATTAAAAA
TTTAGATATTTATGGAACATAATATGTTTGAAACAATAAGACAAAATTATTATTATTATT
ATTATTTTTACTGTTATAATTATGTTGTCTCTTCAATGATTCATAAATAGTTGGACTTGA
TTTTTAAAATGTTTATAATATGATTAGCATAGTTAAATAAAAAAAGTTGAAAAATTAAAA
AAAAACATATAAACACAAATGATGTTTTTTCCTTCAATTTCGATTGATAATTCCTGCAGC
CCAGCTTAATTCTTTTCGAGCTCTTTATGCTTAAGTTTACAATTTAATATTCATACTTTA
AGTATTTTTTGTAGTATCCTAGATATTGTGCTTTAAATGCTCACCCCTCAAAGCACCAGT
AATATTTTCATCCACTGAAATACCATTAAATTTTCAAAAAAATACTATGCATATAATGTT
ATACATATAAACATAAAACGCCATGTAAATCAAAAAATATATAAAAATATGTATAAAAAT
AAATATGCACTAAATATAAGCTAATTATGCATAAAAATTAAAGTGCCCTTTATTAACTAG
AACTAGTCGTAATTATTTATATTTCTATGTTATAAAAAAATCCTCATATAATAATATAAT
TAATATATGTAATGTTTTTTTTATTTTATAATTTTAATATAAAATAATATGTAAATTAAT
TCAAAAAATAAATATAATTGTTGTGAAACAAAAAACGTAATTTTTTCATTTGCCTTCAAA
ATTTAAATTTATTTTAATATTTCCTAAAATATATATACTTTGTGTATAAATATATAAAAA
TATATATTTGCTTATAAATAAATAAAAAATTTTATAAAACATAGGGGGATCCATGGTTGG
TTCGCTAAACTGCATCGTCGCTGTGTCCCAGAACATGGGCATCGGCAAGAACGGGGACCT
GCCCTGGCCACCGCTCAGGAACGAATTTAGATATTTCCAGAGAATGACCACAACCTCTTC
AGTAGAAGGTAAACAGAATCTGGTGATTATGGGTAAGAAGACCTGGTTCTCCATTCCTGA
GAAGAATCGACCTTTAAAGGGTAGAATTAATTTAGTTCTCAGCAGAGAACTCAAGGAACC
TCCACAAGGAGCTCATTTTCTTTCCAGAAGTCTAGATGATGCCTTAAAACTTACTGAACA
ACCAGAATTAGCAAATAAAGTAGACATGGTCTGGATAGTTGGTGGCAGTTCTGTTTATAA
GGAAGCCATGAATCACCCAGGCCATCTTAAACTATTTGTGACAAGGATCATGCAAGACTT
TGAAAGTGACACGTTTTTTCCAGAAATTGATTTGGAGAAATATAAACTTCTGCCAGAATA
CCCAGGTGTTCTCTCTGATGTCCAGGAGGAGAAAGGCATTAAGTACAAATTTGAAGTATA
TGAGAAGAATGATGCTAGCGGAGGAGGTGGATCTGGTGGAGGTGGAAGTGCTAGCGTGAC
AGGGGGAATGGCAAGCAAGTGGGATCAGAAGGGTATGGACATTGCCTATGAGGAGGCGGC
CTTAGGTTACAAAGAGGGTGGTGTTCCTATTGGCGGATGTCTTATCAATAACAAAGACGG
AAGTGTTCTCGGTCGTGGTCACAACATGAGATTTCAAAAGGGATCCGCCACACTACATGG
TGAGATCTCCACTTTGGAAAACTGTGGGAGATTAGAGGGCAAAGTGTACAAAGATACCAC
TTTGTATACGACGCTGTCTCCATGCGACATGTGTACAGGTGCCATCATCATGTATGGTAT
TCCACGCTGTGTTGTCGGTGAGAACGTTAATTTCAAAAGTAAGGGCGAGAAATATTTACA
AACTAGAGGTCACGAGGTTGTTGTTGTTGACGATGAGAGGTGTAAAAAGATCATGAAACA
ATTTATCGATGAAAGACCTCAGGATTGGTTTGAAGATATTGGTGAGGCTTCGGAACCATT
TAAGAACGTCTACTTGCTACCTCAAACAAACCAATTGCTGGGTTTGTACACCATCATCAG
AAATAAGAATACAACTAGACCTGATTTCATTTTCTACTCCGATAGAATCATCAGATTGTT
GGTTGAAGAAGGTTTGAACCATCTACCTGTGCAAAAGCAAATTGTGGAAACTGACACCAA
CGAAAACTTCGAAGGTGTCTCATTCATGGGTAAAATCTGTGGTGTTTCCATTGTCAGAGC
TGGTGAATCGATGGAGCAAGGATTAAGAGACTGTTGTAGGTCTGTGCGTATCGGTAAAAT
TTTAATTCAAAGGGACGAGGAGACTGCTTTACCAAAGTTATTCTACGAAAAATTACCAGA
GGATATATCTGAAAGGTATGTCTTCCTATTAGACCCAATGCTGGCCACCGGTGGTAGTGC
TATCATGGCTACAGAAGTCTTGATTAAGAGAGGTGTTAAGCCAGAGAGAATTTACTTCTT
AAACCTAATCTGTAGTAAGGAAGGGATTGAAAAATACCATGCCGCCTTCCCAGAGGTCAG
AATTGTTACTGGTGCCCTCGACAGAGGTCTAGATGAAAACAAGTATCTAGTTCCAGGGTT
GGGTGACTTTGGTGACAGATACTACTGTGTTTAACTCGATCCCGTTTTTCTTACTTATAT
ATTTATACCAATTGATTGTATTTATAACTGTAAAAATGTGTATGTTGTGTGCATATTTTT
TTTTGTGCATGCACATGCATGTAAATAGCTAAAATTATGAACATTTTATTTTTTGTTCAG
AAAAAAAAAACTTTACACACATAAAATGGCTAGTATGAATAGCCATATTTTATATAAATT
AAATCCTATGAATTTATGACCATATTAAAAATTTAGATATTTATGGAACATAATATGTTT
GAAACAATAAGACAAAATTATTATTATTATTATTATTTTTACTGTTATAATTATGTTGTC
TCTTCAATGATTCATAAATAGTTGGACTTGATTTTTAAAATGTTTATAATATGATTAGCA
TAGTTAAATAAAAAAAGTTGAAAAATTAAAAAAAAACATATAAACACAAATGATGTTTTT
TCCTTCAATTTCGGGTACGTACCCTCGAGGCTAGCGAT
Restriction sites to linearize plasmid Asp718, XbaI, ScaI
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) procedure5-fluorocytosine (5-FC)
Additional remarks genetic modificationTwo general gene targeting DNA constructs pL0001 or pL0037 were used to generate constructs to disrupt the genes selected from the proteome analyses. These constructs are aimed at targeted gene disruption by double cross-over homologous recombination. Sequences of the open reading frame (ORF) and UTR regions of the selected genes were retrieved from PlasmoDB and GeneDB (www.plasmodb.org; www.genedb.org). To replace the ORF of the target genes with the selection cassette containing the pyrimethamine-resistant dhfr/ts of Toxoplasma gondii, the 5′ and 3′ flanking regions of the ORF were cloned up- and down-stream of the selection cassette of pL0001 or pL0037.
To obtain a drug-selectable marker free Δsmac parasites line, the positive T. gondii dhfr/ts selectable marker of pL1351 was replaced with the drug selection cassette of pL0037 (HindIII/EcoRI fragment) that contains a fusion of the positive selectable marker, hdhfr, and the negative selectable marker yfcu (Braks et al., 2006). This construct, pL1386, was used to transfect parasites of reference line wt-GFP-Lucschiz and positive selection with pyrimethamine followed by cloning of parasites resulted in the generation of line 1242cl5 (Δsmac3). To select for parasites which had removed the selectable marker cassette we applied negative selection with 5-fluorocytosine (5-FC)
Additional remarks selection procedureThe mutant does not contain a drug selectable marker which has been removed by the negative selection procedure using the negative selectable marker yfcu (yeast cytosine deaminase and uridyl-phosphoribosyltransferase) and the drug 5-fluorocytosine (5-FC)( (Braks et al., 2006, Nucleic Acids Res 34, e39).
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 1GCCGGTACCGTGTATATATAATCAAATACCAAGG
Additional information primer 13423: 5'forward (Asp718)
Sequence Primer 2GCCAAGCTTCTTGTCCAATGATTTTCCATTG
Additional information primer 23424: 5'reverse (HindIII)
Sequence Primer 3GCCGAATTCCTAAGATCATTATGGGATCATCTG
Additional information primer 33425: 3'forward (EcoRI)
Sequence Primer 4GCCTCTAGAGGGTTGTATTATTAATATTTTATGGG
Additional information primer 43426: 3'reverse (XbaI)
Sequence Primer 5
Additional information primer 5
Sequence Primer 6
Additional information primer 6

  Tagged: Mutant parasite with a tagged gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_1365500
Gene Model P. falciparum ortholog Not available
Gene productintra-erythrocytic P. berghei-induced structures protein 1 exported protein IBIS1
Gene product: Alternative namePlasmodium exported protein IBIS1
Details of the genetic modification
Name of the tagmCherry
Details of taggingC-terminal
Additional remarks: tagging
Commercial source of tag-antibodiesClontech
Type of plasmid/constructPlasmid single 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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CTAAATTGTAAGCGTTAATATTTTGTTAAAATTCGCGTTAAATTTTTGTTAAATCAGCTC
ATTTTTTAACCAATAGGCCGAAATCGGCAAAATCCCTTATAAATCAAAAGAATAGACCGA
GATAGGGTTGAGTGTTGTTCCAGTTTGGAACAAGAGTCCACTATTAAAGAACGTGGACTC
CAACGTCAAAGGGCGAAAAACCGTCTATCAGGGCGATGGCCCACTACGTGAACCATCACC
CTAATCAAGTTTTTTGGGGTCGAGGTGCCGTAAAGCACTAAATCGGAACCCTAAAGGGAG
CCCCCGATTTAGAGCTTGACGGGGAAAGCCGGCGAACGTGGCGAGAAAGGAAGGGAAGAA
AGCGAAAGGAGCGGGCGCTAGGGCGCTGGCAAGTGTAGCGGTCACGCTGCGCGTAACCAC
CACACCCGCCGCGCTTAATGCGCCGCTACAGGGCGCGTCCCATTCGCCATTCAGGCTGCG
CAACTGTTGGGAAGGGCGATCGGTGCGGGCCTCTTCGCTATTACGCCAGCTGGCGAAAGG
GGGATGTGCTGCAAGGCGATTAAGTTGGGTAACGCCAGGGTTTTCCCAGTCACGACGTTG
TAAAACGACGGCCAGTGAGCGCGCGTAATACGACTCACTATAGGGCGAATTGGAGCTCCA
CCGCGGTGGCGGCCGCTCTAGAACTAGTATGGCTCGTAATTGCGAATGCAAAAAGATAGA
TAATGATGATATGGTATCATCCACGAGATATTCCAACAAATTTGGCGAGAAATTTAATTT
GATTTCTTGTACAAAATTGTTTGCGCTGGGCATGTTATTTTTGATATGCAAAAATTGTGA
CAATGTACGAAAAAACAATAAATAAAATAATAGTGACACATTCTAATATATTTTGTTTAT
ATTTTTTTCTTTTTTGTGTTATATGCACATATACATATATTTGATGATACATAATTTTAG
TTATAAGAAAATTTGTTATTTTTATTTTAAAATTTACTATACCCTTTAATTACAACTTTA
TTGTCATTTATTTACTTATTTGTATTTTTACTTTTTAGAGCACGCAAACTACATCCGCAT
ACCAAGAATACCAATATAATGGTTTGATTTTAGGAAAAAGCAGAATATTATCAGAGTTGG
ATCAAAATAAAGATCAAAATTTAGGTTATAAAACACAAAGTTATGAAGATTCTTCGACTG
AAAATGCATCAACCTCATATATGTCAACATTAAAAACTGATGAAGAAAGTAATACGGCGG
GCGAAGATAGAAAAGAAGATAACAATGATGCAGCATTCATTGGAAAAAAATCAACTGAAA
AAAAACCTCACATTGATAGAATATCTTCCACTTTAAATCCAGAATCCACTGACAGAAGAA
AATCTTCTGATGAACAAAAGTCTTCTAACGATAAAAACATATTTAAAGATGTTGATGATC
TAATAAATGGCATCAAATCACGTTATCAAGACATCACCAATAAAATAAAATCACCTGAAT
TCCAAAACGAATGTAAAGGTTATATAAATGTCGCAAAAGAAATGATTGAAGATCATAGAA
ACCATGCTATGAGATTTGTATCTAGAAATTTAAATTCTTTAGGTATTGATCAAATATTCA
ACGATGAGCATGGTGGCTATGCGTTCCTTGCAAAAATTATGTTAACAAAAGTCTTTGTTG
ATAATTTGTTCGTCCCAAATTTCTTAAAAAATAACTCAACAATACTTTCAACTATAGTTT
ACTTTTTAATAATAAGTTTCATCGTAAGCAACTATCTTGATGCCACTCAAAATAATGAAA
GAGAAAGAAAAAATTTGAATAAATCTAACATATTTTGTAGAGCAAAACCTATGGGATCCG
GTGGAGGTGGAAGATCCATGGTGAGCAAGGGCGAGGAGGATAACATGGCCATCATCAAGG
AGTTCATGCGCTTCAAGGTGCACATGGAGGGCTCCGTGAACGGCCACGAGTTCGAGATCG
AGGGCGAGGGCGAGGGCCGCCCCTACGAGGGCACCCAGACCGCCAAGCTGAAGGTGACCA
AGGGTGGCCCCCTGCCCTTCGCCTGGGACATCCTGTCCCCTCAGTTCATGTACGGCTCCA
AGGCCTACGTGAAGCACCCCGCCGACATCCCCGACTACTTGAAGCTGTCCTTCCCCGAGG
GCTTCAAGTGGGAGCGCGTGATGAACTTCGAGGACGGCGGCGTGGTGACCGTGACCCAGG
ACTCCTCCCTGCAGGACGGCGAGTTCATCTACAAGGTGAAGCTGCGCGGCACCAACTTCC
CCTCCGACGGCCCCGTAATGCAGAAGAAGACCATGGGCTGGGAGGCCTCCTCCGAGCGGA
TGTACCCCGAGGACGGCGCCCTGAAGGGCGAGATCAAGCAGAGGCTGAAGCTGAAGGACG
GCGGCCACTACGACGCTGAGGTCAAGACCACCTACAAGGCCAAGAAGCCCGTGCAGCTGC
CCGGCGCCTACAACGTCAACATCAAGTTGGACATCACCTCCCACAACGAGGACTACACCA
TCGTGGAACAGTACGAACGCGCCGAGGGCCGCCACTCCACCGGCGGCATGGACGAGCTGT
ACAAGTAATCTAGAATTATTAATATATATGAATATATATACATCGTTGTATGCGTAATAT
ACGTGTAACTGTTTTTTTTAATTTTTATATTAACTTTTCCTTATACATTTTAATGCCATT
AACTAATAATATTTTACTTGAAAAAATTCTAAATTTTTATAACTCTAATATAAAATCTGT
GAATAATAAAAAATTAGCATAAAGTACGAATTAAACATCGAATATTTGTTTTAAAGGGGC
ACACATTATGTGCATATAATTTTATTCTTTGAATTTAATTTTTTGCTTATAATTAGATTA
AATTAATATAATTTTTTTATTGAAAATAAAAACTTATTTTAAAAAATTAACAAATGGAAG
TTAATGAAAAATACGAAAAAAGTAAAGGGTTAATTCTTATATGGTCGGTACCGAGCTCGA
ATTAATTCGATATCGAAATTGAAGGAAAAAACATCATTTGTGTTTATATGTTTTTTTTTA
ATTTTTCAACTTTTTTTATTTAACTATGCTAATCATATTATAAACATTTTAAAAATCAAG
TCCAACTATTTATGAATCATTGAAGAGACAACATAATTATAACAGTAAAAATAATAATAA
TAATAATAATTTTGTCTTATTGTTTCAAACATATTATGTTCCATAAATATCTAAATTTTT
AATATGGTCATAAATTCATAGGATTTAATTTATATAAAATATGGCTATTCATACTAGCCA
TTTTATGTGTGTAAAGTTTTTTTTTTCTGAACAAAAAATAAAATGTTCATAATTTTAGCT
ATTTACATGCATGTGCATGCACAAAAAAAAATATGCACACAACATACACATTTTTACAGT
TATAAATACAATCAATTGGTATAAATATATAAGTAAGAAAAACGGGATCAAAGCTAGCTT
CTGTATTTCCGCTAGACAGCCATCTCCATCTGGATTCGTCCGTGCGGGACGTAGCCCACG
ACCTCAAAATCCTCGGCGGTGAAATCGTCGATTTCCTTGATGCGTTCCTTGTTGAGGATG
TTCACAATGGGGAACGGTCTCGGTTCTCTCCGCAGCTGCTCTTTTAAAGCCTCGACATGG
TTCGTGTAGACATGCGTGTTCCCCATGAAGTGAATGAACTCCTTAGGTTTTAGGTTGCAG
ACGTGTGCAACCATGAGCGTCAAAAGCGAATAGGAAGCGATGTTGAAGGGGACGCCGAGG
CCGACATCGCACGACCGCTGATACATGATGCACGACAGCTCCTTCTGGTCGTTCACGTAG
AACTGGCACAACAAGTGACAAGGCGGCAGCGCCATTTCGTCCAGCGCTGCAGGATTCCAG
GCAGTCATGAGCATGCGACGATCTGTTGGATTCGTTCTCAGCATCTGGATCACATTCTTC
AGCTGGTCGACGCCCTGCCCTGTGTAGTCTGTGTGCATGTCTTTGTATGCCGCGCCGAAG
TGTCTCCACTGGAAGCCGTAGCCCGGGCCGATGTCTCCGACCTCTCGGTGGGGGAGATTG
CGCGAATCGAGGAACTCGCGTGTCACATTCTTGTCCCAGATCTTCACGCCCTTCTCAGAA
AGATGGTTTGCGTTCGTGTCGCCGCGAATGAACCACAGCAACTCTTCGAGGACCCCTTTC
CAGAACACACGCTTTGTGGTGAGAAGTGGAAAGGCCTGATCCAGCGAGTAGCGCATAGTG
CAGCCGAATTTGGAGATGACACCAACGCCCGTTCGGTCATCCATTGTCCTTCCATTGTTA
ATAATGTCGGCAATGAGATCAAGGTACTGGAACTCTTCATGGCCTCTAAAGTGAACATGC
GGAACGGCCCGAATCAGTTCCTTTTGCTCGCGTTTTTTCCGGTCTTCTTCGTCCATCCAC
GCCAACACCGGGGCAATGGCTGCGGCCGAAGAAGGAGCCTGCAACCCGTGCACGGGAGTT
GTCTCCCTCGTGGACGTCAAGGAGCTCATTGCGTTGCTCGGTTCCGCAGTGGCTGCGTCG
TCAGTCTTCCTTCTCTTCTCGAGAACCACAAAGTCGTAGGGAACCCCGTTGTCTGAGAAG
GTCTTGGAAATGAAGATGGGTCGATACGTCGCTTCATTGTCCTTCTCTCTTCCGAGCTCC
GGACAAAAGGGAACGAACACAGACTCGGCAGGAGCTGCAGCCTGCGCAGCAGTTGATTTG
TTTGAAAGAATGTCATCTCCGGGGAACGCAGGGAAGAAAACGTCGCACGGAAACTCGCGG
GCTACACGCGTGATGTACAGGTGAGAGGCAACGCCCAGAGACAGCGCTGCCTCGTACAGT
CCCGCTCCTCCCACGACAAAAATCTGGTCGACAGAATCCTTGTACTCTTCCTCCAGAAGG
CTGAGAGCTGCTGGGAGTGAAGCACAGACTCGGACGCGCTGCTGGCCTTCAGCTTGAGGC
TTCTCCGCCGCAATGTCTTCTTCTTTGAGGGAAGAGGAAACGACGATGTTCAATCTGTCC
ACGAGGGGTCTAAACTTTCGAGGCATGCTTTCCCAGGTTTTCCGTCCCATGACAACGGCG
TTGAATCTCTTGCCGACTGATGGAGAGGGAAGTCCAGAGTCGCCCGTCTTTGCAAATTTC
CTGGGAAGCCACCCGTTCAGGCGACTGGCTTCTTCGGGCGTCGTTTTTGTCACACGAGAA
AAGTGTTTGAAATCTGTGGTCAAGTGGGGCCACGGGAGGCCGTTGTTGATGCCGATGCCC
CTCTTGGGGGTCATCGCGACGACCAGACACACCGGTTTATGCATTGATCCTTTGTAACAT
TTAGGTGTGTATTTATATATATAAGCAATATACAAATGAAAATACTATTATAAAACGGAA
ACTCAAAAAATGTTGAAACAAATAGTCAATTTTTAAAATGAATGTGTAAATTGTTTAAGT
TTTATGACAAAAAAAGGAAAAGTGTATATAAAATAAAAATTGGAAAAAATTTATATATTA
CATGCCATATAATACCATTTATTTACTATTTATATCTTAATATAAAAAATTGTAGCAAAA
ATATATAATCGTATTTTATATATTTAATATTATATAACCCCTTAAAAAAAAAAATTTAAT
TGAAAAAATTTTAATTTATGCACTTTTTTATTGTATTATATATATTCCGTTATTTTAATT
ACACATAAACTTAATAGAAAAATTAATTCGCTGCTAAATAAAAATTATAGGGCTATATAT
GATGGGATCATATTCTATCTTTCTCTTTTTCAAATYCTTTATATTTTAGAATACAAATCG
CTTTAAACAATAAAAAAGAGGTATATCCTACCCGAATTGACACACACAATAAATAATAAA
TATATAAATAATAAAAATATAAATATATAAATTATAAATATATAAATTATATATATAAAT
ATATTAACATGTGTGTAATTTGTTAAAATTGATATTAAACAGCTTTGCATGGGGAAAGAG
TGGCTTTTATAAGCGAATCATATAATACAGTGTTAATTTTAATATATAAAATATAATTTC
CTTAAATTGTTATTTAACATTTTTAATTGCCTACCCARGGTAATATAATATCCAAATGAC
AATCAAAGATAAATGAAATACCGCTCCATTTTTCCTATATATCCATTTTCATTTTGTGTA
AATGTTCATAAAATTTTATCATATACTCATAAAAAACAGGGGAAAAATGAAAAAATGAAC
GTCCACCCCGTGTGAATATGCTCATTTTGTATACTATACATTTTTTTTTATATAATTTAT
ATTTTCCAAATAAAAAAAATGTTTGTACATGTATATATCAATATTGTGTTATGGCATGAT
GAAAAACAATTATTTTATTCGTTTTTAAATTTGTAGAATTTCCCTAAATTATTAATGATT
TCATATTATAATATATGTATCTATATATTTTTAATGTGTAATTCAAAATGGCGAAAAAGA
AACAAACCATACTTTATAAGCAACTCTCTTTGAGCACTTACCATAAAAAATTATACTACT
AAATTCAAATTAAATAAAAAAATATATATTATAAAATTGGCTTTAATTATTTATAAGCAC
TTGCTTTGTTTTACTCAAAAAGTATTGGCATAAGTATAAACATAAAACCAATTTTTCACA
AAATATATTATATGTCCATACAACTATATCCGAACYTTCAAAACACATCCAAATTTCCAT
TTACCACATGCAAGCAAAAGGATATTAAATAAAAAAAAATTATTTATATTTTCATGATCA
CTTAAATAATATTTAAAAAAAAAAGAGACCATATATAAAATGTATAATTTAAATATAGCT
AACTTATTAAAAAAAATGAATATTGAGCATATATCCATAATTTAAAACACTTCTATTTTA
TGTTATTTGATATATAAGTCGATAAATTATTGCATGTATGCATATAGACAGATTAATATA
CACCAAAATAATAAAAATTATATATAGTTCATATATTATTCTGAATAATAAATTAGCTTT
CCTCTTAATTCACTTGGTATGTATATATATATATATATATATATATATATATATATGGAT
AAATAAATATAAGGTCTATAACAGGAAAAAATGTGTATTAACTTTTTTTTATAATTATTT
TATATTTTTTCTCCATTTATTTAAATATGTTAAAATTATTTCCCATTTTATAAACAAAAA
TTAATCCATAAAATAGCAAAATAACAAAATGACAAAATAACAAAAACAAAAAAACGCATT
ATATGAGTTCATTTTACACAATCCAAAGAAAAAGAAAATATGCTTAAAATATTCATAACA
CACTTTTAAGCTATAAAAATATATATATTTATTTGTGTCTATATTACCAACTCAATTTAA
TAGATGTGTTATGTGATTAATTCATACACAAACATACAAAAATAAACACCATTTTTGAAA
AGATTAATTTGAAATCATATATGTTATTATTATATATTTTTATATCAATGAAATTTAAAA
AAATAAAAGGGAAATCAATGTATTAAAAATAATTATATGTTATTTTATTTCCACAATATT
TATTTATTATTTATTGTTGACCTGCAGGCATGCAAGCTAGCTTATCGATACCGTCGACCT
CGAGGGGGGGCCCGGTACCCAGCTTTTGTTCCCTTTAGTGAGGGTTAATTGCGCGCTTGG
CGTAATCATGGTCATAGCTGTTTCCTGTGTGAAATTGTTATCCGCTCACAATTCCACACA
ACATACGAGCCGGAAGCATAAAGTGTAAAGCCTGGGGTGCCTAATGAGTGAGCTAACTCA
CATTAATTGCGTTGCGCTCACTGCCCGCTTTCCAGTCGGGAAACCTGTCGTGCCAGCTGC
ATTAATGAATCGGCCAACGCGCGGGGAGAGGCGGTTTGCGTATTGGGCGCTCTTCCGCTT
CCTCGCTCACTGACTCGCTGCGCTCGGTCGTTCGGCTGCGGCGAGCGGTATCAGCTCACT
CAAAGGCGGTAATACGGTTATCCACAGAATCAGGGGATAACGCAGGAAAGAACATGTGAG
CAAAAGGCCAGCAAAAGGCCAGGAACCGTAAAAAGGCCGCGTTGCTGGCGTTTTTCCATA
GGCTCCGCCCCCCTGACGAGCATCACAAAAATCGACGCTCAAGTCAGAGGTGGCGAAACC
CGACAGGACTATAAAGATACCAGGCGTTTCCCCCTGGAAGCTCCCTCGTGCGCTCTCCTG
TTCCGACCCTGCCGCTTACCGGATACCTGTCCGCCTTTCTCCCTTCGGGAAGCGTGGCGC
TTTCTCATAGCTCACGCTGTAGGTATCTCAGTTCGGTGTAGGTCGTTCGCTCCAAGCTGG
GCTGTGTGCACGAACCCCCCGTTCAGCCCGACCGCTGCGCCTTATCCGGTAACTATCGTC
TTGAGTCCAACCCGGTAAGACACGACTTATCGCCACTGGCAGCAGCCACTGGTAACAGGA
TTAGCAGAGCGAGGTATGTAGGCGGTGCTACAGAGTTCTTGAAGTGGTGGCCTAACTACG
GCTACACTAGAAGGACAGTATTTGGTATCTGCGCTCTGCTGAAGCCAGTTACCTTCGGAA
AAAGAGTTGGTAGCTCTTGATCCGGCAAACAAACCACCGCTGGTAGCGGTGGTTTTTTTG
TTTGCAAGCAGCAGATTACGCGCAGAAAAAAAGGATCTCAAGAAGATCCTTTGATCTTTT
CTACGGGGTCTGACGCTCAGTGGAACGAAAACTCACGTTAAGGGATTTTGGTCATGAGAT
TATCAAAAAGGATCTTCACCTAGATCCTTTTAAATTAAAAATGAAGTTTTAAATCAATCT
AAAGTATATATGAGTAAACTTGGTCTGACAGTTACCAATGCTTAATCAGTGAGGCACCTA
TCTCAGCGATCTGTCTATTTCGTTCATCCATAGTTGCCTGACTCCCCGTCGTGTAGATAA
CTACGATACGGGAGGGCTTACCATCTGGCCCCAGTGCTGCAATGATACCGCGAGACCCAC
GCTCACCGGCTCCAGATTTATCAGCAATAAACCAGCCAGCCGGAAGGGCCGAGCGCAGAA
GTGGTCCTGCAACTTTATCCGCCTCCATCCAGTCTATTAATTGTTGCCGGGAAGCTAGAG
TAAGTAGTTCGCCAGTTAATAGTTTGCGCAACGTTGTTGCCATTGCTACAGGCATCGTGG
TGTCACGCTCGTCGTTTGGTATGGCTTCATTCAGCTCCGGTTCCCAACGATCAAGGCGAG
TTACATGATCCCCCATGTTGTGCAAAAAAGCGGTTAGCTCCTTCGGTCCTCCGATCGTTG
TCAGAAGTAAGTTGGCCGCAGTGTTATCACTCATGGTTATGGCAGCACTGCATAATTCTC
TTACTGTCATGCCATCCGTAAGATGCTTTTCTGTGACTGGTGAGTACTCAACCAAGTCAT
TCTGAGAATAGTGTATGCGGCGACCGAGTTGCTCTTGCCCGGCGTCAATACGGGATAATA
CCGCGCCACATAGCAGAACTTTAAAAGTGCTCATCATTGGAAAACGTTCTTCGGGGCGAA
AACTCTCAAGGATCTTACCGCTGTTGAGATCCAGTTCGATGTAACCCACTCGTGCACCCA
ACTGATCTTCAGCATCTTTTACTTTCACCAGCGTTTCTGGGTGAGCAAAAACAGGAAGGC
AAAATGCCGCAAAAAAGGGAATAAGGGCGACACGGAAATGTTGAATACTCATACTCTTCC
TTTTTCAATATTATTGAAGCATTTATCAGGGTTATTGTCTCATGAGCGGATACATATTTG
AATGTATTTAGAAAAATAAACAAATAGGGGTTCCGCGCACATTTCCCCGAAAAGTGCCAC
Restriction sites to linearize plasmid EcoRI
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 1AACTAGTATGGCTCGTAATTGCGAATG
Additional information primer 1L6288 (SpeI); PBANKA_136550 Forward
Sequence Primer 2AGGATCCCATAGGTTTTGCTCTACAAAATATG
Additional information primer 2L6289 (BamHI); PBANKA_136550 Reverse
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

  Transgene: Mutant parasite expressing a transgene
Type and details of transgene
Is the transgene Plasmodium derived Transgene: not Plasmodium
Transgene nameA fusion of GFP (gfp-mu3) and Luciferase Firefly (LucIAV)
Details of the genetic modification
Inducable system usedNo
Additional remarks inducable system
Type of plasmid/constructPlasmid 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
ATTGTGAGCGGATAACAATTTCACACAGGAAACAGCTATGACCATGATTACGCCAAGCTT
CCGCGGGTATATGGTAAAGAACCTACTAACACAATAAAATATTTAAATAATGTATTTCCT
ATAAATAAATTTACAGATTTATTTTTTAATACAAAAGATATAGATATACCAGAAATAAAT
GATCAGTTTAAAGGTTTTAAATTCTTTATGACATCATTTATAAATCATGGATCATATCCA
CTAACAATAGAATGTGGTGTAACAAATGGTGGAACTAGTTATAAAAGAGCAATTATTTTA
TTGCATGTTCGAACTGATTTAAAAGATAGACCAGTTTCATTTTGTGATTTTCGAAAAGGA
GAATTATATAATTATTTGAATGCTTATACTGAAGGGGATGTATGCATAATAATTTCCAAA
TCAAATACAAGTTTTGGTTTTAGATGCCCAGTAAATACAAAAAAAATGCCAAAAAATTGT
TTTACGCAAGTATATGAAAAAGGGTATCTAAATGACGCCAATAAAATTAATACTAAAAAT
GTTATTAACTATTCATTTGAAAATCCAGAATATGCGCTAGCTGGTTYTAATTATACATTA
ACAAAATCGTATCAATTTGAATGTCATTGTGTAGATAAAGAAACAGAACAAATTGTAAAA
ACGGTTTTAGTCAAATATGTAAATGAAGATGAAATATATGATTATAATGATTTTCCAATG
GTGAATCACAAACCTATTATTGCACATCCAAATAAAACACATCAAGCTTGCATGCCTGCA
GGAATTCGATATCGAATTCCCATGGAACCAAAATAACATATTAATGAATTTTCTTTTATT
GTGAAAATGTTCTTATTTGTTTTAAAAAATCAAAGTAAATTTAATTCCTTTAATTGTGCT
AAATATTGTGTTTGTATAACTATAATGACATTTCCTATGCTTTTTTAATATTATATATAA
TTTTTTGTGTTTTTTCAAAAAAAACGTGTATTTGTTTTTATTTGAGTGTACAATTTGCAT
AGTGAGTTTAAAGTGTTTGAAAATTATTTAATTTATAATTTTCATAAAACGTTTAATCAT
TTATCAAAATTGCGGTTGCAATATAGACAGAAAATATTAACTTAATTTTCATTGTTTTGT
AAAACATTTTCATTATTTTTTTTAACAAAATTAAAGTAAATGAGATGGTAAATTATTTTA
ATTATAACCATTTGTTTAATTTTTTTTATTTTATTACTATTATTTTTTTTCTTTTGTGCT
GCGCAATTAAAAAGAGAATTAAGGGTGCACCAATATATTTAAGCAATCTTTTGCGACACA
CAAGAAAAAATATAACATCAACATATTTAAAATAATGGAAATTTTTTAATAATTTATTCT
ATATATATTTAATATATGTAAAAAAACTAATATTGTAATATAATTAAATAAAACTTGAAA
TTGATATAATATGTATATATATTGCATGTTTAGCTTTAAATAATATTTAAGTATTGATTT
TCTTTACACACAAAAACATTTGCCTTTTTATGTGAGGATTATTGTTTCTTTATTTATATT
GCTATGAATTTATTCTTAGTATAAATTTATTTTTTTTCGGCTTTGAAAATAATTTTATAT
ATATAAAAAGAAAAAATATGGAAAATAAAATTCATAAATCATATTTGACACATTTCAATT
TTTTTTTCTTATTTCACAGCTAGCCATATAATAATATATATGTGTCTATGGGTTCAAATA
GCTTAATTTCAATGTGCGTGATTTTTTTTTATAAAAAAACTAATTTTATGTTTGGTTATG
CATGCAACTGTTATAAAATAAAAATTGTAAAATTTGTTTTTTTGTACACATATTTGTACT
TTTTTTAATGCACATGAAAAAAGAAAGGAAAAAAAAGGGGGAAAAAAAAGGAAAAAAAAA
GGGAAAAAAGGGAAAAAACGTACATCTACGCATTGTTATTTAGCAATAAAATAGAGAACA
AATCTATAAATATATAATGTGTACAACAAATTAATTTAATTTATATTTTTACTGTAATTT
CATAATAATTTTCATTTTTACATTTTAATGTTTTTTTAATTGGAATATAATTCAAAATGA
TTTTGACACATTTTACTTATAGAAGAAGGCTATATAATTATATAAGTGTGTATATATAAG
TATTGTATGGTAATTGTTTTATAAATTAAATTTTTAAAAAACATTATTCTATTTTATAAT
TTGTAAATTAAAAATATTAATAAAATAAAACGATATAAAAGGATCCATGAGTAAAGGAGA
AGAACTTTTCACTGGAGTTGTCCCAATTCTTGTTGAATTAGATGGTGATGTTAATGGGCA
CAAATTTTCTGTCAGTGGAGAGGGTGAAGGTGATGCAACATACGGAAAACTTACCCTTAA
ATTTATTTGCACTACTGGAAAACTACCTGTTCCATGGCCAACACTTGTCACTACTTTCGG
TTATGGTGTTCAATGCTTTGCGAGATACCCAGATCATATGAAACAGCATGACTTTTTCAA
GAGTGCCATGCCCGAAGGTTATGTACAGGAAAGAACTATATTTTTCAAAGATGACGGGAA
CTACAAGACACGTGCTGAAGTCAAGTTTGAAGGTGATACCCTTGTTAATAGAATCGAGTT
AAAAGGTATTGATTTTAAAGAAGATGGAAACATTCTTGGACACAAATTGGAATACAACTA
TAACTCACACAATGTATACATCATGGCAGACAAACAAAAGAATGGAATCAAAGTTAACTT
CAAAATTAGACACAACATTGAAGATGGAAGCGTTCAACTAGCAGACCATTATCAACAAAA
TACTCCAATTGGCGATGGCCCTGTCCTTTTACCAGACAACCATTACCTGTCCACACAATC
TGCCCTTTCGAAAGATCCCAACGAAAAGAGAGACCACATGGTCCTTCTTGAGTTTGTAAC
AGCTGCTGGGATTACACATGGCATGGATGAACTATACAAAGGGATCCTGGCTAGCCAGTC
GACCTGCAGGCATGCAAGCTTGCGGCCGATCCAAATGGAAGACGCCAAAAACATAAAGAA
AGGCCCGGCGCCATTCTATCCGCTGGAAGATGGAACCGCTGGAGAGCAACTGCATAAGGC
TATGAAGAGATACGCCCTGGTTCCTGGAACAATTGCTTTTACAGATGCACATATCGAGGT
GAACATCACGTACGCGGAATACTTCGAAATGTCCGTTCGGTTGGCAGAAGCTATGAAACG
ATATGGGCTGAATACAAATCACAGAATCGTCGTATGCAGTGAAAACTCTCTTCAATTCTT
TATGCCGGTGTTGGGCGCGTTATTTATCGGAGTTGCAGTTGCGCCCGCGAACGACATTTA
TAATGAACGTGAATTGCTCAACAGTATGAACATTTCGCAGCCTACCGTAGTGTTTGTTTC
CAAAAAGGGGTTGCAAAAAATTTTGAACGTGCAAAAAAAATTACCAATAATCCAGAAAAT
TATTATCATGGATTCTAAAACGGATTACCAGGGATTTCAGTCGATGTACACGTTCGTCAC
ATCTCATCTACCTCCCGGTTTTAATGAATACGATTTTGTACCAGAGTCCTTTGATCGTGA
CAAAACAATTGCACTGATAATGAATTCCTCTGGATCTACTGGGTTACCTAAGGGTGTGGC
CCTTCCGCATAGAACTGCCTGCGTCAGATTCTCGCATGCCAGAGATCCTATTTTTGGCAA
TCAAATCATTCCGGATACTGCGATTTTAAGTGTTGTTCCATTCCATCACGGTTTTGGAAT
GTTTACTACACTCGGATATTTGATATGTGGATTTCGAGTCGTCTTAATGTATAGATTTGA
AGAAGAGCTGTTTTTACGATCCCTTCAGGATTACAAAATTCAAAGTGCGTTGCTAGTACC
AACCCTATTTTCATTCTTCGCCAAAAGCACTCTGATTGACAAATACGATTTATCTAATTT
ACACGAAATTGCTTCTGGGGGCGCACCTCTTTCGAAAGAAGTCGGGGAAGCGGTTGCAAA
ACGCTTCCATCTTCCAGGGATACGACAAGGATATGGGCTCACTGAGACTACATCAGCTAT
TCTGATTACACCCGAGGGGGATGATAAACCGGGCGCGGTCGGTAAAGTTGTTCCATTTTT
TGAAGCGAAGGTTGTGGATCTGGATACCGGGAAAACGCTGGGCGTTAATCAGAGAGGCGA
ATTATGTGTCAGAGGACCTATGATTATGTCCGGTTATGTAAACAATCCGGAAGCGACCAA
CGCCTTGATTGACAAGGATGGATGGCTACATTCTGGAGACATAGCTTACTGGGACGAAGA
CGAACACTTCTTCATAGTTGACCGCTTGAAGTCTTTAATTAAATACAAAGGATATCAGGT
GGCCCCCGCTGAATTGGAATCGATATTGTTACAACACCCCAACATCTTCGACGCGGGCGT
GGCAGGTCTTCCCGACGATGACGCCGGTGAACTTCCCGCCGCCGTTGTTGTTTTGGAGCA
CGGAAAGACGATGACGGAAAAAGAGATCGTGGATTACGTCGCCAGTCAAGTAACAACCGC
GAAAAAGTTGCGCGGAGGAGTTGTGTTTGTGGACGAAGTACCGAAAGGTCTTACCGGAAA
ACTCGACGCAAGAAAAATCAGAGAGATCCTCATAAAGGCCAAGAAGGGCGGAAAGATCGC
CGTGTAATTCTAGAAGATCCCGTTTTTCTTACTTATATATTTATACCAATTGATTGTATT
TATAACTGTAAAAATGTGTATGTTGTGTGCATATTTTTTTTTGTGCATGCACATGCATGT
AAATAGCTAAAATTATGAACATTTTATTTTTTGTTCAGAAAAAAAAAACTTTACACACAT
AAAATGGCTAGTATGAATAGCCATATTTTATATAAATTAAATCCTATGAATTTATGACCA
TATTAAAAATTTAGATATTTATGGAACATAATATGTTTGAAACAATAAGACAAAATTATT
ATTATTATTATTATTTTTACTGTTATAATTATGTTGTCTCTTCAATGATTCATAAATAGT
TGGACTTGATTTTTAAAATGTTTATAATATGATTAGCATAGTTAAATAAAAAAAGTTGAA
AAATTAAAAAAAAACATATAAACACAAATGATGTTTTTTCCTTCAATTTCGGGTACCGAG
CTCGAATTCTCTTGAGCCCGTTAATGAAATAGATACAATTCATTCATGTTATATACATCT
AGAACATAATCTGAATATGGTTCAAGTTAAATGTCCAAAAATTATAAAAAGTGATGATAT
TTTTGATGGTAATACCATAATAGACACCAAGGTAACATCACGAAGTAGTCAACAAAATAA
TTTTTATTTAGAAAATACAGATGTTGAACCAGAAGAAATAGAGAAATATAAAAATATAGA
ATACATACCAGAAAACGATGAAGTAATGCATCTAGACAAAAAAGAAAAGCTAGATGATAT
ATTACCAGGTGTTATCATATTAGATAAAAATAAAATGTTCAAAGAAAAAGGACATTTCAC
TTTTGTTACTCCATTAATTGTAGAAAAGGTATTAATATTAAAAATATATTGTGATAATAC
TAAAACAATAATTAATAATATGAAAGGGAAAAAAGGTATTACAGTAATAAGGATTTCTCA
AAATACAACAAAAAATAAATTTTATGGATGTGACTTTTCAGGTAATTCTAAAAAAACATT
TTACTATTCCAATGTTTATGATTTAGAAAAAAAAAATGAGTTTTGTGAAATAGAATTAAA
AGAAAATATAGTAGTTAGCTTAAATTGTCCAACTGGTAAAATTAATCCAAAAAATTGTTT
TAGAAATGTATATATAAAAAGTAATATGAATGAACAAACAACCGAAAATATAGAAAATAT
ATTTAACGAAATAAAAGTTATAGATGCAGATTATTTTATAAATAATTCATCAACCTTTTT
GATGATTTCCAAAATTACAAAAAAAGAGTTTGATTTTTATTGTACATGTGAAGATTATAA
AACCAAAAATATAGGAACAATATATATTAAAAATTATGAATATCTAGATTCAAAACCTAA
ATATAAAAATAAACAAATTTCCTATATAGATGTAGTTCCATACCCGCGGGGAAAGGGCG
Restriction sites to linearize plasmid KspI (SacII)
Selectable marker used to select the mutant parasitegfp (FACS)
Promoter of the selectable markerama-1
Selection (positive) procedureFACS (flowsorting)
Selection (negative) procedureNo
Additional remarks genetic modificationThe GFP-Luciferase gene (1 copy) has been inserted into the 230p locus (PB000214.00.0) by double cross-over integration
Additional remarks selection procedureThis reporter mutant expressing GFP-Luciferase does not contain a drug-selectable marker. This mutant has been selected by FACS sorting after transfection based on GFP fluorescence
Other details transgene
Promoter
Gene Model of Parasite PBANKA_0915000
Gene Model P. falciparum ortholog PF3D7_1133400
Gene productapical membrane antigen 1
Gene product: Alternative nameAMA-1
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
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 PBANKA_0306000
Gene product6-cysteine protein
Gene product: Alternative name230p
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