Summary

RMgm-122
Malaria parasiteP. berghei
Genotype
Transgene
Transgene not Plasmodium: GFP (gfp-mut3)
Promoter: Gene model: PBANKA_1300700; Gene model (P.falciparum): PF3D7_1475500; Gene product: LCCL domain-containing protein (LAP2, LCCL/lectin adhesive-like protein 2)
3'UTR: Gene model: PBANKA_0719300; Gene product: bifunctional dihydrofolate reductase-thymidylate synthase, putative (dhfr/ts)
Insertion locus: Gene model: Not available; Gene product: Not available (small subunit ribosomal rna gene (c- or d-type unit))
Phenotype Gametocyte/Gamete; Oocyst; Sporozoite;
Last modified: 18 February 2009, 13:21
  *RMgm-122
Successful modificationThe parasite was generated by the genetic modification
The mutant contains the following genetic modification(s) Introduction of a transgene
Reference (PubMed-PMID number) Reference 1 (PMID number) : 15935755
Reference 2 (PMID number) : 18848846
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/ResearcherS.M. Khan, G.R. Mair, C.J. Janse, A.P. Waters
Name Group/DepartmentLeiden Malaria Research Group
Name InstituteLeiden University Medical Center (LUMC)
CityLeiden
CountryThe Netherlands
Name of the mutant parasite
RMgm numberRMgm-122
Principal name480
Alternative namePbCCp1-GFP
Standardized name
Is the mutant parasite cloned after genetic modificationNo
Phenotype
Asexual blood stageNot different from wild type
Gametocyte/GameteGFP expression in female gametocytes. No expression could be detected in asexual blood stages and male gametocytes.
Fertilization and ookineteNot tested
OocystNo GFP expression could be detected in oocysts at day 11 and 17
SporozoiteNo GFP expression could be detected in salivary gland sporozoites
Liver stageNot different from wild type
Additional remarks phenotype

Mutant/mutation
The mutant expresses GFP under the control of the promoter of the gene encoding LAP2 (LCCL/lectin adhesive-like protein 2; LCCL domain containing protein CCp1; PF14_0723).

Protein (function)
The lap2 (ccp1) gene is a member of a small conserved gene family, encoding proteins with multiple adhesive domains, for example a Lgl1 (LCCL)-lectin adhesive domain. In P. falciparum the LCCL domain-containing proteins are termed PfCCp's.
See also the phenotype description of mutants lacking expression of LAP2/CCp1 (RMgm-118, RMgm-121).

Phenotype
Phenotype analyses indicate absence of promoter activity in asexual blood stages and male gametocytes and active transcription in female gametocytes. Promoter activity appears to be absent in oocysts and sporozoites. The absence in male gametocytes and prescence in females is in agreement with proteome analysis of males and females (Khan, S.M. et al., 2005, Cell, 121, 675-87). See further 'Additional information.

Additional information
The construct GFPcon (RMgm-5) containing GFP under the control of the eef1a promoter formed the basis for the construct driving expression of GFP under control of the ccp1/lap2 promoter. This constructs contains a c/d-ssurrna target fragment for integration into the c/d-rrna genomic locus. GFP is flanked by the 3'-UTR sequence of the dhfr gene of P. berghei. The eef1a promoter region is replaced by a 1986 bp fragment (EcoRV/BamHI) of the promoter region of ccp1/lap2.
Because it has been shown that 3'-UTR regions can influence expression of genes in Plasmodium, care has to be taken in interpretation of expression of transgenes when 3'-UTR sequences of heterologous genes are used.

Other mutants


  Transgene: Mutant parasite expressing a transgene
Type and details of transgene
Is the transgene Plasmodium derived Transgene: not Plasmodium
Transgene nameGFP (gfp-mut3)
Details of the genetic modification
Inducable system usedNo
Additional remarks inducable system
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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ATCTTTATAAATGAAGCAATACCACATTTTTACACAAATATATTACATTTTTTATGTATT
GCATATGTGAAATGTATGAGTGATATAATATGGTAGACATGAATAGGCTAAAACATAGAT
AAAACCGAACTAAACAAATAAATATAGGAAAATACCCCAATACGAATATTTGTAACATTA
AAAATTAAAAAGACACACTTTAATTAAATAAAATAGAGACGTGAATAAAAATGTATTTAT
AATAAAAGTACGTAAAAAGAAAGCGTTACCAAAACATAAATGATTTTTTTCTAACTAAAT
AAAATTGCATATGTGAAGGTATAATATTTTCCTACATAAAATGGCGTTTATTGGTTTTTT
AATATGTTTTACTTCATTTTGATTTATTACACATATATAAATTTTTTACAAAATTACGCC
ATTAATATAGACATTTTTAAGAATGTTGTATTTATTGATCATATTCGAACTAAATTCCGT
TGTAAATTCACTCAGGTGCATGCAAAAGATGCGGTATTTTTTTGTACTATTAAAAATTTA
TATATACAATAATTGTTCAAAATACATAATTTTTTTTAAGCATTTTACAAACACAAATAA
TTTTTTCAAAATAAAATAAATTGTTACGTATCTCGAAAATAGTGTATTTAATATTATACA
AATGTAAGGGCAATACATATATATATATTTATTTATTTAAAAGACAATTTATAAATGCAT
ATTATTCATAAATTTTTATAATATTTCTCGATAATTTTGTTCGCAATTTGATGGTGACCC
CTTATACATTTGAATTTGCTTTAAAAATATTAAAAAGGAAAATTTTTTTTTGTTATATAG
AGTCATCATGCTTGATTATTTTATTTCTGAATTTAATTTATCCTCATGTATTTATTATAA
CAATACGACGTGAAGTTAAAACACCAGTTGCATTGATCAATTTAAAATTTTGATAAAAAG
TATTTAAAAATAAAAGGTTTATAAATAGGGGCTTATTCATTTATATTTCTTTTTATTATA
TTTTCTTTATATTTTTCAACAAATATTTATAATTCGTAATTTTTTCGTCAATTTAATATA
TTTGCCTTTAAGTTTTTCGTAAATTACACTTCAGTAATAATAATAAAGCTACATTTTATA
GTGCGCATTGTTAAGCATATAATCGATAGGACTTTTAATATATATTTTGTTTTTACACAA
AATAAGAAGAAGGTCCCTACATTTTTTTCGTTAAAAAGGAAATCTCAATAATATTAAATT
GTGTGTAATTTTTTTTGATGTATACAAATATGACGTGTATATACGCAAAGGTGCTTGTAA
AACAAATGGTATTAAGCTGTGTGGTCACATTTGTAGTTACATAGCAATTATATATATATC
ATTTCTAATTTTAATTCAATTTAATTTCGGTATTAACTATGCATATAAACATATTGTTAT
TATTATCTTGCCTATCATAATAAATTTTGTGTCTATATAGAAATGTATTATCTATTTTTT
AGCGAAGTAGCATAAGTATTATTTATATTCAAAGAAATAAATAAAGGTATTAAAATTAAT
ACGAAAATAAAAATTAAAGGATGATGGCGATTATTAGTTCTTTTTAATTCCATCATCTAT
CCTTATATGTAGTATATTGGGGTTAGAGATAGTGGGTTATGAATACCAAAAATATAGAAA
ATTGCAAAGGAGTTGATTTATTCTCTGTTGTATAAGAATGTACACAGTTAAAGTTAGTAT
ATATGTGTATATTTTTCAGTCTATATAAATATACTTTTGTTTTCTCATTGGTATGTTCAA
TTTTTTATATTATTTTACGTGTCGATGGATACAATTTAGTATTTTTTTTAATTTTGTATT
TTTGAATTATATTAATCTAGATTATTCATATATTTTTTCATATGCAGATAGAAAAAGGTT
TCATACGTTAGTTTATCCTACAAATTGAATATTAACACATACATATATTTATATTTTTTA
TTAATAGAAGGATCCATGAGTAAAGGAGAAGAACTTTTCACTGGAGTTGTCCCAATTCTT
GTTGAATTAGATGGTGATGTTAATGGGCACAAATTTTCTGTCAGTGGAGAGGGTGAAGGT
GATGCAACATACGGAAAACTTACCCTTAAATTTATTTGCACTACTGGAAAACTACCTGTT
CCATGGCCAACACTTGTCACTACTTTCGGTTATGGTGTTCAATGCTTTGCGAGATACCCA
GATCATATGAAACAGCATGACTTTTTCAAGAGTGCCATGCCCGAAGGTTATGTACAGGAA
AGAACTATATTTTTCAAAGATGACGGGAACTACAAGACACGTGCTGAAGTCAAGTTTGAA
GGTGATACCCTTGTTAATAGAATCGAGTTAAAAGGTATTGATTTTAAAGAAGATGGAAAC
ATTCTTGGACACAAATTGGAATACAACTATAACTCACACAATGTATACATCATGGCAGAC
AAACAAAAGAATGGAATCAAAGTTAACTTCAAAATTAGACACAACATTGAAGATGGAAGC
GTTCAACTAGCAGACCATTATCAACAAAATACTCCAATTGGCGATGGCCCTGTCCTTTTA
CCAGACAACCATTACCTGTCCACACAATCTGCCCTTTCGAAAGATCCCAACGAAAAGAGA
GACCACATGGTCCTTCTTGAGTTTGTAACAGCTGCTGGGATTACACATGGCATGGATGAA
CTATACAAATAAATTCTAGAAGATCCCGTTTTTCTTACTTATATATTTATACCAATTGAT
TGTATTTATAACTGTAAAAATGTGTATGTTGTGTGCATATTTTTTTTTGTGCATGCACAT
GCATGTAAATAGCTAAAATTATGAACATTTTATTTTTTGTTCAGAAAAAAAAAACTTTAC
ACACATAAAATGGCTAGTATGAATAGCCATATTTTATATAAATTAAATCCTATGAATTTA
TGACCATATTAAAAATTTAGATATTTATGGAACATAATATGTTTGAAACAATAAGACAAA
ATTATTATTATTATTATTATTTTTACTGTTATAATTATGTTGTCTCTTCAATGATTCATA
AATAGTTGGACTTGATTTTTAAAATGTTTATAATATGATTAGCATAGTTAAATAAAAAAA
GTTGAAAAATTAAAAAAAAACATATAAACACAAATGATGTTTTTTCCTTCAATTTCGGGT
ACCGAGCTCGAATTCAACCTGGTTGATCTTGCCAGTAGTCATATGCTTGTCTCAAAGATT
AAGCCATGCAAGTGAAAGTATATGCACATTTATTGCAGAAACTGCGAACGGCTCATTAAA
ACAGTTATAATCTACTTGACATTTTATTATAAGGATAACTACGGAAAAGCTGTAGCTAAT
ACTTGTTAAGTACTTTTACTCCCCGGAGTAATTGTATGTATTTGTTAAGACCCCTAAGAA
AAAATGATATTAAAGGAATTATAACAAAGAAGCAACACATAATATAATTATTCAGTGTGT
ATCAATCGAGTTTCTGACCTATCAGCTTTTGATGTTAGGGTATTGACCTAACATGGCTTT
GACGGGTAACGGGGAATTAGAGTTCGATTCCGGAGAGGGAGCCTGAGAAATAGCTACCAC
ATCTAAGGAAGGCAGCAGGCGCGTAAATTACCCAATTCTAAATAAGAGAGGTAGTGACAA
GAAATAACAATATAAGGCCAAATTTTGGTTTTATAATTGGAATGATGGGAATTTAAAACC
TTCCCAAAAATCAATTGGAGGGCAAGTCTGGTGCCAGCAGCCGCGGTAATTCCAGCTCCA
ATAGCGTATATTAAAATTGTTGCAGTTAAAACGCTCGTAGTTGAACTTCAAGGGTATAAT
TATTTTAAGCAACTCACTTGGAAAGAATCATGACTTCTGTCACTGCTTTTATCCTTGTTG
CAGTTCTTTTAATACAGGGCCCTTTGAGAGCCCATTAATTTATGACTGGGTTTCTCGTTA
CTTTGAGTAAATTAGAGTGTTTAAAGCAAACAGATAAAGCGTATTTTACTGTGTTTGAAT
ACTATAGCATGGAATAACAACATTGAATAGGTCAAAAGTTTTTGAAAAATTTTTCTTATT
TTGGCTTAGATACAGTTAATAGGAGTAGCTTGGGGGCATTTGTATTCAGATGTCAGAGGT
GAAATTCTTAGATTTTCTGGAGACAAACAACTGCGAAAGCATTTGCCTAAAATACTTCCA
TTAATCAAGAACGAAAGTTAAGGGAGTGAAGACGATCAGATACCGTCGTAATCTTAACCA
TAAACTATGCCGACTAAGTGTTGGATGAAAATTTATAAATAAAACTATCTTCTTTAAAGG
AGTAGTTTTTTAGATGCTTCCTTCAGTACCTTATGAGAAATCAAAGTCTTTGGGTTCTGG
GGCGAGTATTCGCGCAAGCGAGAAAGTTAAAAGAATTGACGGAAGGGCACCACCAGGCGT
GGAGCTTGCGGCTTAATTTGACTCAACACGGGGAAACTCACTAGTTTAAGACAAGAGTAG
GATTGACAGATTAATAGCTCTTTCTTGATTTCTTGGATGGTGATGCATGGCCGTTTTTAG
TTCGTGAATATGATTTGTCTGGTTAATTCCGATAACGAACGAGATCTTAACCTGCTAATT
AGCGGTGGATATGTGATATTCTTCGAAGGTGGACTAACTATAGCGTTTTCGAAGGTATGT
TGCATAATCAAATTGGTTTACCCTTTGTTTTTTTGTAGCATATTCTTTTATTTCGTTGGG
TTTTTTCCCTAGTAAGGATGTATCTGCTTTATTTAATGCTTCTTAGAGGAACGATGTGTG
TCTAACACAAGGAAGTTTAAGGCAACAACAGGTCTGTGATGTCCTTAGATATACTAGGCT
GCACGCGTGCTACACTGATATGTAAAACGAGTATTTAAAATTATATCTGTATGGTAGATA
ATTTAATTTCTACGTATTATCAGCATATACTTTTCCTACACTGAAATAGTGAAGGTAATC
TTTATCAATACATATCGTGATGGGGATAGATTATTGCAATTATTAATCTTGAACGAGGAA
TGCCTAGTAAGCATGATTCATCAGATTGTGCTGACTACGTCCCTGCCCTTTGTACACACC
GCCCGTCGCTCCTACCGATTGAAAGATATGATGAATTGTTTGGACAAGAAAATAGAAATT
TTATTTTTATTTTTTTTGGAAGGACCGTAAATCCTATCTTTTAAAGGAAGGAGAAGTCGT
AACAAGGTTTCCGTAGGTGAATTCACTGGCCGTCGTTTTACAACGTCGTGACTGGGAAAA
CCCTGGCGTTACCCAACTTAATCGCCTTGCAGCACATCCCCCTTTCGCCAGCTGGCGTAA
TAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGTTGCGCAGCCTGAATGGCGAATG
GCGCCTGATGCGGTATTTTCTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTG
CACTCTCAGTACAATCTGCTCTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAAC
ACCCGCTGACGCGCCCTGACGGGCTTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGT
GACCGTCTCCGGGAGCTGCATGTGTCAGAGGTTTTCACCGTCATCACCGAAACGCGCGAG
ACGAAAGGGCCTCGTGATACGCCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTC
TTAGACGTCAGGTGGCACTTTTCGGGGAAATGTGCGCGGAACCCCTATTTGTTTATTTTT
CTAAATACATTCAAATATGTATCCGCTCATGAGACAATAACCCTGATAAATGCTTCAATA
ATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCCGTGTCGCCCTTATTCCCTTTTT
TGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAGTAAAAGATGC
TGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAAGAT
CCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCACTTTTAAAGTTCTGCT
ATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTCGCCGCATACA
CTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTACGGATGG
CATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACACTGCGGCCAA
CTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACATGGG
GGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGA
CGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATTAACTGG
CGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGT
TGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGG
AGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCCTC
CCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACA
GATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTC
ATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGAT
CCTTTTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTC
AGACCCCGTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTG
CTGCTTGCAAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCT
ACCAACTCTTTTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTCCT
TCTAGTGTAGCCGTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCT
CGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGG
GTTGGACTCAAGACGATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTC
GTGCACACAGCCCAGCTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGA
GCATTGAGAAAGCGCCACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGG
CAGGGTCGGAACAGGAGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTA
TAGTCCTGTCGGGTTTCGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGG
GGGGCGGAGCCTATGGAAAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTG
CTGGCCTTTTGCTCACATGTTCTTTCCTGCGTTATCCCCTGATTCTGTGGATAACCGTAT
TACCGCCTTTGAGTGAGCTGATACCGCTCGCCGCAGCCGAACGACCGAGCGCAGCGAGTC
AGTGAGCGAGGAAGCGGAAGAGCGCCCAATACGCAAACCGCCTCTCCCCGCGCGTTGGCC
GATTCATTAATGCAGCTGGCACGACAGGTTTCCCGACTGGAAAGCGGGCAGTGAGCGCAA
CGCAATTAATGTGAGTTAGCTCACTCATTAGGCACCCCAGGCTTTACACTTTATGCTTCC
GGCTCGTATGTTGTGTGGAATTGTGAGCGGATAACAATTTCACACAGGAAACAGCTATGA
CCATGATTACGCCAAGCTTGCATGCCTGCAGGTCAACAATAAATAATAAATAAATATTGT
GGAAATAAAATAACATATAATTATTTTTAATACATTGATTTCCCTTTTATTTTTTTAAAT
TTCATTGATATAAAAATATATAATAATAACATATATGATTTCAAATTAATCTTTTCAAAA
ATGGTGTTTATTTTTGTATGTTTGTGTATGAATTAATCACATAACACATCTATTAAATTG
AGTTGGTAATATAGACACAAATAAATATATATATTTTTATAGCTTAAAAGTGTGTTATGA
ATATTTTAAGCATATTTTCTTTTTCTTTGGATTGTGTAAAATGAACTCATATAATGCGTT
TTTTTGTTTTTGTTATTTTGTCATTTTGTTATTTTGCTATTTTATGGATTAATTTTTGTT
TATAAAATGGGAAATAATTTTAACATATTTAAATAAATGGAGAAAAAATATAAAATAATT
ATAAAAAAAAGTTAATACACATTTTTTCCTGTTATAGACCTTATATTTATTTATCCATAT
ATATATATATATATATATATATATATATATACATACCAAGTGAATTAAGAGGAAAGCTAA
TTTATTATTCAGAATAATATATGAACTATATATAATTTTTATTATTTTGGTGTATATTAA
TCTGTCTATATGCATACATGCAATAATTTATCGACTTATATATCAAATAACATAAAATAG
AAGTGTTTTAAATTATGGATATATGCTCAATATTCATTTTTTTTAATAAGTTAGCTATAT
TTAAATTATACATTTTATATATGGTCTCTTTTTTTTTTAAATATTATTTAAGTGATCATG
AAAATATAAATAATTTTTTTTTATTTAATATCCTTTTGCTTGCATGTGGTAAATGGAAAT
TTGGATGTGTTTTGAARGTTCGGATATAGTTGTATGGACATATAATATATTTTGTGAAAA
ATTGGTTTTATGTTTATACTTATGCCAATACTTTTTGAGTAAAACAAAGCAAGTGCTTAT
AAATAATTAAAGCCAATTTTATAATATATATTTTTTTATTTAATTTGAATTTAGTAGTAT
AATTTTTTATGGTAAGTGCTCAAAGAGAGTTGCTTATAAAGTATGGTTTGTTTCTTTTTC
GCCATTTTGAATTACACATTAAAAATATATAGATACATATATTATAATATGAAATCATTA
ATAATTTAGGGAAATTCTACAAATTTAAAAACGAATAAAATAATTGTTTTTCATCATGCC
ATAACACAATATTGATATATACATGTACAAACATTTTTTTTATTTGGAAAATATAAATTA
TATAAAAAAAAATGTATAGTATACAAAATGAGCATATTCACACGGGGTGGACGTTCATTT
TTTCATTTTTCCCCTGTTTTTTATGAGTATATGATAAAATTTTATGAACATTTACACAAA
ATGAAAATGGATATATAGGAAAAATGGAGCGGTATTTCATTTATCTTTGATTGTCATTTG
GATATTATATTACCYTGGGTAGGCAATTAAAAATGTTAAATAACAATTTAAGGAAATTAT
ATTTTATATATTAAAATTAACACTGTATTATATGATTCGCTTATAAAAGCCACTCTTTCC
CCATGCAAAGCTGTTTAATATCAATTTTAACAAATTACACACATGTTAATATATTTATAT
ATATAATTTATATATTTATAATTTATATATTTATATTTTTATTATTTATATATTTATTAT
TTATTGTGTGTGTCAATTCGGGTAGGATATACCTCTTTTTTATTGTTTAAAGCGATTTGT
ATTCTAAAATATAAAGRATTTGAAAAAGAGAAAGATAGAATATGATCCCATCATATATAG
CCCTATAATTTTTATTTAGCAGCGAATTAATTTTTCTATTAAGTTTATGTGTAATTAAAA
TAACGGAATATATATAATACAATAAAAAAGTGCATAAATTAAAATTTTTTCAATTAAATT
TTTTTTTTTAAGGGGTTATATAATATTAAATATATAAAATACGATTATATATTTTTGCTA
CAATTTTTTATATTAAGATATAAATAGTAAATAAATGGTATTATATGGCATGTAATATAT
AAATTTTTTCCAATTTTTATTTTATATACACTTTTCCTTTTTTTGTCATAAAACTTAAAC
AATTTACACATTCATTTTAAAAATTGACTATTTGTTTCAACATTTTTTGAGTTTCCGTTT
TATAATAGTATTTTCATTTGTATATTGCTTATATATATAAATACACACCTAAATGTTACA
AAGGATCAATGCATAAACCGGTGTGTCTGGTCGTCGCGATGACCCCCAAGAGGGGCATCG
GCATCAACAACGGCCTCCCGTGGCCCCACTTGACCACAGATTTCAAACACTTTTCTCGTG
TGACAAAAACGACGCCCGAAGAAGCCAGTCGCCTGAACGGGTGGCTTCCCAGGAAATTTG
CAAAGACGGGCGACTCTGGACTTCCCTCTCCATCAGTCGGCAAGAGATTCAACGCCGTTG
TCATGGGACGGAAAACCTGGGAAAGCATGCCTCGAAAGTTTAGACCCCTCGTGGACAGAT
TGAACATCGTCGTTTCCTCTTCCCTCAAAGAAGAAGACATTGCGGCGGAGAAGCCTCAAG
CTGAAGGCCAGCAGCGCGTCCGAGTCTGTGCTTCACTCCCAGCAGCTCTCAGCCTTCTGG
AGGAAGAGTACAAGGATTCTGTCGACCAGATTTTTGTCGTGGGAGGAGCGGGACTGTACG
AGGCAGCGCTGTCTCTGGGCGTTGCCTCTCACCTGTACATCACGCGTGTAGCCCGCGAGT
TTCCGTGCGACGTTTTCTTCCCTGCGTTCCCCGGAGATGACATTCTTTCAAACAAATCAA
CTGCTGCGCAGGCTGCAGCTCCTGCCGAGTCTGTGTTCGTTCCCTTTTGTCCGGAGCTCG
GAAGAGAGAAGGACAATGAAGCGACGTATCGACCCATCTTCATTTCCAAGACCTTCTCAG
ACAACGGGGTTCCCTACGACTTTGTGGTTCTCGAGAAGAGAAGGAAGACTGACGACGCAG
CCACTGCGGAACCGAGCAACGCAATGAGCTCCTTGACGTCCACGAGGGAGACAACTCCCG
TGCACGGGTTGCAGGCTCCTTCTTCGGCCGCAGCCATTGCCCCGGTGTTGGCGTGGATGG
ACGAAGAAGACCGGAAAAAACGCGAGCAAAAGGAACTGATTCGGGCCGTTCCGCATGTTC
ACTTTAGAGGCCATGAAGAGTTCCAGTACCTTGATCTCATTGCCGACATTATTAACAATG
GAAGGACAATGGATGACCGAACGGGCGTTGGTGTCATCTCCAAATTCGGCTGCACTATGC
GCTACTCGCTGGATCAGGCCTTTCCACTTCTCACCACAAAGCGTGTGTTCTGGAAAGGGG
TCCTCGAAGAGTTGCTGTGGTTCATTCGCGGCGACACGAACGCAAACCATCTTTCTGAGA
AGGGCGTGAAGATCTGGGACAAGAATGTGACACGCGAGTTCCTCGATTCGCGCAATCTCC
CCCACCGAGAGGTCGGAGACATCGGCCCGGGCTACGGCTTCCAGTGGAGACACTTCGGCG
CGGCATACAAAGACATGCACACAGACTACACAGGGCAGGGCGTCGACCAGCTGAAGAATG
TGATCCAGATGCTGAGAACGAATCCAACAGATCGTCGCATGCTCATGACTGCCTGGAATC
CTGCAGCGCTGGACGAAATGGCGCTGCCGCCTTGTCACTTGTTGTGCCAGTTCTACGTGA
ACGACCAGAAGGAGCTGTCGTGCATCATGTATCAGCGGTCGTGCGATGTCGGCCTCGGCG
TCCCCTTCAACATCGCTTCCTATTCGCTTTTGACGCTCATGGTTGCACACGTCTGCAACC
TAAAACCTAAGGAGTTCATTCACTTCATGGGGAACACGCATGTCTACACGAACCATGTCG
AGGCTTTAAAAGAGCAGCTGCGGAGAGAACCGAGACCGTTCCCCATTGTGAACATCCTCA
ACAAGGAACGCATCAAGGAAATCGACGATTTCACCGCCGAGGATTTTGAGGTCGTGGGCT
ACGTCCCGCACGGACGAATCCAGATGGAGATGGCTGTCTAGCGGAAATACAGAAGCTAGC
TTTGATCCCGTTTTTCTTACTTATATATTTATACCAATTGATTGTATTTATAACTGTAAA
AATGTGTATGTTGTGTGCATATTTTTTTTTGTGCATGCACATGCATGTAAATAGCTAAAA
TTATGAACATTTTATTTTTTGTTCAGAAAAAAAAAACTTTACACACATAAAATGGCTAGT
ATGAATAGCCATATTTTATATAAATTAAATCCTATGAATTTATGACCATATTAAAAATTT
AGATATTTATGGAACATAATATGTTTGAAACAATAAGACAAAATTATTATTATTATTATT
ATTTTTACTGTTATAATTATGTTGTCTCTTCAATGATTCATAAATAGTTGGACTTGATTT
TTAAAATGTTTATAATATGATTAGCATAGTTAAATAAAAAAAGTTGAAAAATTAAAAAAA
AACATATAAACACAAATGATGTTTTTTCCTTCAATTTCGAT
Restriction sites to linearize plasmid
Selectable marker used to select the mutant parasitetgdhfr
Promoter of the selectable markerpbdhfr
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modificationThe construct GFPcon (RMgm-5) containing GFP under the control of the eef1a promoter formed the basis for the construct driving expression of GFP under control of the ccp1/lap2 promoter.
This constructs contains a c/d-ssurrna target fragment for integration into the c/d-rrna genomic locus. GFP is flanked by the 3'-UTR sequence of the dhfr gene of P. berghei. The eef1a promoter region is replaced by a 1986 bp fragment (EcoRV/BamHI) of the promoter region of ccp1/lap2.

Because it has been shown that 3'-UTR regions can influence expression of genes in Plasmodium, care has to be taken in interpretation of expression of transgenes when 3'-UTR sequences of heterologous genes are used.

The gfp gene has been integrated into the genome by single cross-over integration. Therefore the possibility exists of reversion to the wild-type genotype by removal of the integrated DNA construct.
Additional remarks selection procedure
Other details transgene
Promoter
Gene Model of Parasite PBANKA_1300700
Gene Model P. falciparum ortholog PF3D7_1475500
Gene productLCCL domain-containing protein
Gene product: Alternative nameLAP2, LCCL/lectin adhesive-like protein 2
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 1aaGATATCtttataaatgaagcaataccac
Additional information primer 1forward
Sequence Primer 2ttGGATCCttctattaataaaaaatataaatatatg
Additional information primer 2reverse
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 / InsertionInsertion locus
Gene Model of Parasite Not available
Gene productNot available
Gene product: Alternative namesmall subunit ribosomal rna gene (c- or d-type unit)
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