Summary

RMgm-204
Malaria parasiteP. berghei
Genotype
DisruptedGene model (rodent): PBANKA_0819800; Gene model (P.falciparum): PF3D7_0918900; Gene product: gamma-glutamylcysteine synthetase (γ-GCS)
Phenotype Asexual bloodstage; Fertilization and ookinete; Oocyst; Sporozoite;
Last modified: 4 July 2010, 10:36
  *RMgm-204
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) : 19229315
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 507cl1 (RMgm-7)
Other information parent lineP.berghei ANKA 507cl1 (RMgm-7) is a reference ANKA mutant line which expresses GFP under control of a constitutive promoter (PubMed: PMID: 16242190).
The mutant parasite was generated by
Name PI/ResearcherJ. Vega-Rodriguez, C.J. Janse, A.E. Serrano
Name Group/DepartmentDepartment of Microbiology and Medical Zoology
Name InstituteUniversity of Puerto Rico, School of Medicine
CitySan Juan
CountryPuerto Rico
Name of the mutant parasite
RMgm numberRMgm-204
Principal name866cl1; 985cl1
Alternative namepbggcs−1; pbggcs−2
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Phenotype
Asexual blood stageBlood stages show a small decrease in the growth rate in mice.
Gametocyte/GameteNot different from wild type
Fertilization and ookineteNormal gametocyte production. The ookinete production in Anopheles stephensi mosquitoes was reduced by 30-40% as compared to wild type parasites. Ookinetes had a reduced capacity to develop into (mature) oocysts.
OocystOokinetes had a reduced capacity to develop into (mature) oocysts. At two days after feeding of mosquitoes the number of young oocysts (stained with anti-P28 antibodies) was reduced by 70-90% when compared to wild type. At day 12 after feeding the oocysts numbers were reduced by 65-85%. Day 12 oocysts were significantly smaller than wild type oocysts and no formation of sporozoites was observed.
SporozoiteNo formation of sporozoites
Liver stageNot tested
Additional remarks phenotype

Mutant/mutation
The mutant lacks expression of gamma-glutamylcysteine synthetase (γ-GCS).

Protein (function)
γ-GCS catalyzes the rate limiting step during GSH biosynthesis. GSH is a thiol-based tripeptide implicated in a variety of cellular processes, including detoxification of xenobiotics and protection against reactive oxygen species (see also 'Additional information').

Phenotype
The phenotype analyses indicate that γ-GCS is not essential for blood stage development. Despite a significant reduction in GSH levels, blood stage forms of mutant parasites showed only a minor defect in growth as compared to wild type, resulting in a small growth delay in mice. In contrast, a dramatic effect of the lack of expression of γ-GCS was observed on development of the parasites in the mosquito. Infection of mosquitoes resulted in reduced numbers of stunted oocysts that did not produce sporozoites. These results indicate that de novo synthesis of GSH is pivotal for development in the mosquito.

Additional information
GSH levels have been analysed by HPLC in blood stages of wild type and mutant parasites.  Glutathione levels were significantly reduced (P<0.001) the mutant parasites (0.2-1.0 nmol/109 parasites) as compared to wild type parasites (7.4 nmol/109 parasites).

γ-GCS catalyzes the rate limiting step during GSH biosynthesis. GSH is a thiol-based tripeptide implicated in a variety of cellular processes, including detoxification of xenobiotics and protection against reactive oxygen species.

In most eukaryotic organisms, redox-active enzymes, such as catalase, superoxide dismutase, and peroxidases as well as an enzymatic cascade that generates reduced electron donors, i.e. glutathione (GSH) and thioredoxin (Trx), sustain the cellular redox homeostasis. This redox network is split into two major arms, the GSH and the Trx system, that serve complementary functions in antioxidant defense and DNA synthesis. The malarial parasite Plasmodium lacks two central antioxidant enzymes: (i) catalase that typically detoxifies hydrogen peroxide and (ii) a classical glutathione peroxidase, a selenoenyzme that reduces lipid hydroperoxides to their alcohols. This apparent deficiency raises doubts about the relevance of the glutathione (GSH) pathway in detoxification of oxidative stress in Plasmodium. However, supportive of a role for GSH metabolism in the detoxification process are the observations that the P. falciparum glutathione S-tranferase enzyme, which conjugates GSH to other molecules via the sulfhydryl group, displays peroxidase activity.

Evidence has been presented that P. falciparum does not utilize GSH from the host red blood cell since the parasite membrane is neither permeable to host GSH nor γ-glutamylcysteine. Plasmodium is therefore thought to be dependent on its own GSH biosynthetic pathway. GSH is synthesized in Plasmodium by consecutive reactions facilitated by the enzymes γ-glutamylcysteine synthetase (γ-GCS) and glutathione synthetase (GS). However, it has been hypothesized that host GSH can be transported into the P. berghei food vacuole via hemoglobin-containing endocytic vesicles, based on data showing that GSH can detoxify the toxic ferriprotoporphyrin IX inside the parasite's food vacuole.

See also mutants RMgm-403 and RMgm-404 that lack expression of glutathione reductase (GR). Phenotype analyses of these mutants indicate that similar to what was reported for γ-GCS, GR is not essential for parasite blood stage development but it does play a critical role during oocyst development in the mosquito

Other mutants
RMgm-403, RMgm-404: Mutants lacking expression of glutathione reductase (GR).


  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_0819800
Gene Model P. falciparum ortholog PF3D7_0918900
Gene productgamma-glutamylcysteine synthetase
Gene product: Alternative nameγ-GCS
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
Click to view information
Click to hide information
Plasmid/construct sequence
Click to view information
Click to hide information
AGCTTGCATGCCTGCAGGTCAACAATAAATAATAAATAAATATTGTGGAAATAAAATAAC
ATATAATTATTTTTAATACATTGATTTCCCTTTTATTTTTTTAAATTTCATTGATATAAA
AATATATAATAATAACATATATGATTTCAAATTAATCTTTTCAAAAATGGTGTTTATTTT
TGTATGTTTGTGTATGAATTAATCACATAACACATCTATTAAATTGAGTTGGTAATATAG
ACACAAATAAATATATATATTTTTATAGCTTAAAAGTGTGTTATGAATATTTTAAGCATA
TTTTCTTTTTCTTTGGATTGTGTAAAATGAACTCATATAATGCGTTTTTTTGTTTTTGTT
ATTTTGTCATTTTGTTATTTTGCTATTTTATGGATTAATTTTTGTTTATAAAATGGGAAA
TAATTTTAACATATTTAAATAAATGGAGAAAAAATATAAAATAATTATAAAAAAAAGTTA
ATACACATTTTTTCCTGTTATAGACCTTATATTTATTTATCCATATATATATATATATAT
ATATATATATATATATACATACCAAGTGAATTAAGAGGAAAGCTAATTTATTATTCAGAA
TAATATATGAACTATATATAATTTTTATTATTTTGGTGTATATTAATCTGTCTATATGCA
TACATGCAATAATTTATCGACTTATATATCAAATAACATAAAATAGAAGTGTTTTAAATT
ATGGATATATGCTCAATATTCATTTTTTTTAATAAGTTAGCTATATTTAAATTATACATT
TTATATATGGTCTCTTTTTTTTTTAAATATTATTTAAGTGATCATGAAAATATAAATAAT
TTTTTTTTATTTAATATCCTTTTGCTTGCATGTGGTAAATGGAAATTTGGATGTGTTTTG
AARGTTCGGATATAGTTGTATGGACATATAATATATTTTGTGAAAAATTGGTTTTATGTT
TATACTTATGCCAATACTTTTTGAGTAAAACAAAGCAAGTGCTTATAAATAATTAAAGCC
AATTTTATAATATATATTTTTTTATTTAATTTGAATTTAGTAGTATAATTTTTTATGGTA
AGTGCTCAAAGAGAGTTGCTTATAAAGTATGGTTTGTTTCTTTTTCGCCATTTTGAATTA
CACATTAAAAATATATAGATACATATATTATAATATGAAATCATTAATAATTTAGGGAAA
TTCTACAAATTTAAAAACGAATAAAATAATTGTTTTTCATCATGCCATAACACAATATTG
ATATATACATGTACAAACATTTTTTTTATTTGGAAAATATAAATTATATAAAAAAAAATG
TATAGTATACAAAATGAGCATATTCACACGGGGTGGACGTTCATTTTTTCATTTTTCCCC
TGTTTTTTATGAGTATATGATAAAATTTTATGAACATTTACACAAAATGAAAATGGATAT
ATAGGAAAAATGGAGCGGTATTTCATTTATCTTTGATTGTCATTTGGATATTATATTACC
YTGGGTAGGCAATTAAAAATGTTAAATAACAATTTAAGGAAATTATATTTTATATATTAA
AATTAACACTGTATTATATGATTCGCTTATAAAAGCCACTCTTTCCCCATGCAAAGCTGT
TTAATATCAATTTTAACAAATTACACACATGTTAATATATTTATATATATAATTTATATA
TTTATAATTTATATATTTATATTTTTATTATTTATATATTTATTATTTATTGTGTGTGTC
AATTCGGGTAGGATATACCTCTTTTTTATTGTTTAAAGCGATTTGTATTCTAAAATATAA
AGRATTTGAAAAAGAGAAAGATAGAATATGATCCCATCATATATAGCCCTATAATTTTTA
TTTAGCAGCGAATTAATTTTTCTATTAAGTTTATGTGTAATTAAAATAACGGAATATATA
TAATACAATAAAAAAGTGCATAAATTAAAATTTTTTCAATTAAATTTTTTTTTTTAAGGG
GTTATATAATATTAAATATATAAAATACGATTATATATTTTTGCTACAATTTTTTATATT
AAGATATAAATAGTAAATAAATGGTATTATATGGCATGTAATATATAAATTTTTTCCAAT
TTTTATTTTATATACACTTTTCCTTTTTTTGTCATAAAACTTAAACAATTTACACATTCA
TTTTAAAAATTGACTATTTGTTTCAACATTTTTTGAGTTTCCGTTTTATAATAGTATTTT
CATTTGTATATTGCTTATATATATAAATACACACCTAAATGTTACAAAGGATCAATGCAT
AAACCGGTGTGTCTGGTCGTCGCGATGACCCCCAAGAGGGGCATCGGCATCAACAACGGC
CTCCCGTGGCCCCACTTGACCACAGATTTCAAACACTTTTCTCGTGTGACAAAAACGACG
CCCGAAGAAGCCAGTCGCCTGAACGGGTGGCTTCCCAGGAAATTTGCAAAGACGGGCGAC
TCTGGACTTCCCTCTCCATCAGTCGGCAAGAGATTCAACGCCGTTGTCATGGGACGGAAA
ACCTGGGAAAGCATGCCTCGAAAGTTTAGACCCCTCGTGGACAGATTGAACATCGTCGTT
TCCTCTTCCCTCAAAGAAGAAGACATTGCGGCGGAGAAGCCTCAAGCTGAAGGCCAGCAG
CGCGTCCGAGTCTGTGCTTCACTCCCAGCAGCTCTCAGCCTTCTGGAGGAAGAGTACAAG
GATTCTGTCGACCAGATTTTTGTCGTGGGAGGAGCGGGACTGTACGAGGCAGCGCTGTCT
CTGGGCGTTGCCTCTCACCTGTACATCACGCGTGTAGCCCGCGAGTTTCCGTGCGACGTT
TTCTTCCCTGCGTTCCCCGGAGATGACATTCTTTCAAACAAATCAACTGCTGCGCAGGCT
GCAGCTCCTGCCGAGTCTGTGTTCGTTCCCTTTTGTCCGGAGCTCGGAAGAGAGAAGGAC
AATGAAGCGACGTATCGACCCATCTTCATTTCCAAGACCTTCTCAGACAACGGGGTTCCC
TACGACTTTGTGGTTCTCGAGAAGAGAAGGAAGACTGACGACGCAGCCACTGCGGAACCG
AGCAACGCAATGAGCTCCTTGACGTCCACGAGGGAGACAACTCCCGTGCACGGGTTGCAG
GCTCCTTCTTCGGCCGCAGCCATTGCCCCGGTGTTGGCGTGGATGGACGAAGAAGACCGG
AAAAAACGCGAGCAAAAGGAACTGATTCGGGCCGTTCCGCATGTTCACTTTAGAGGCCAT
GAAGAGTTCCAGTACCTTGATCTCATTGCCGACATTATTAACAATGGAAGGACAATGGAT
GACCGAACGGGCGTTGGTGTCATCTCCAAATTCGGCTGCACTATGCGCTACTCGCTGGAT
CAGGCCTTTCCACTTCTCACCACAAAGCGTGTGTTCTGGAAAGGGGTCCTCGAAGAGTTG
CTGTGGTTCATTCGCGGCGACACGAACGCAAACCATCTTTCTGAGAAGGGCGTGAAGATC
TGGGACAAGAATGTGACACGCGAGTTCCTCGATTCGCGCAATCTCCCCCACCGAGAGGTC
GGAGACATCGGCCCGGGCTACGGCTTCCAGTGGAGACACTTCGGCGCGGCATACAAAGAC
ATGCACACAGACTACACAGGGCAGGGCGTCGACCAGCTGAAGAATGTGATCCAGATGCTG
AGAACGAATCCAACAGATCGTCGCATGCTCATGACTGCCTGGAATCCTGCAGCGCTGGAC
GAAATGGCGCTGCCGCCTTGTCACTTGTTGTGCCAGTTCTACGTGAACGACCAGAAGGAG
CTGTCGTGCATCATGTATCAGCGGTCGTGCGATGTCGGCCTCGGCGTCCCCTTCAACATC
GCTTCCTATTCGCTTTTGACGCTCATGGTTGCACACGTCTGCAACCTAAAACCTAAGGAG
TTCATTCACTTCATGGGGAACACGCATGTCTACACGAACCATGTCGAGGCTTTAAAAGAG
CAGCTGCGGAGAGAACCGAGACCGTTCCCCATTGTGAACATCCTCAACAAGGAACGCATC
AAGGAAATCGACGATTTCACCGCCGAGGATTTTGAGGTCGTGGGCTACGTCCCGCACGGA
CGAATCCAGATGGAGATGGCTGTCTAGCGGAAATACAGAAGCTAGCTTTGATCCCGTTTT
TCTTACTTATATATTTATACCAATTGATTGTATTTATAACTGTAAAAATGTGTATGTTGT
GTGCATATTTTTTTTTGTGCATGCACATGCATGTAAATAGCTAAAATTATGAACATTTTA
TTTTTTGTTCAGAAAAAAAAAACTTTACACACATAAAATGGCTAGTATGAATAGCCATAT
TTTATATAAATTAAATCCTATGAATTTATGACCATATTAAAAATTTAGATATTTATGGAA
CATAATATGTTTGAAACAATAAGACAAAATTATTATTATTATTATTATTTTTACTGTTAT
AATTATGTTGTCTCTTCAATGATTCATAAATAGTTGGACTTGATTTTTAAAATGTTTATA
ATATGATTAGCATAGTTAAATAAAAAAAGTTGAAAAATTAAAAAAAAACATATAAACACA
AATGATGTTTTTTCCTTCAATTTCGATGACCCCCATAAAGAAAGAATAAAAAATAATTTA
AGACAAATTAGTAAAAGACAATATTTAAAATCTTTAGCTTATAAATTTATTTCTGGGGAA
GACTACACTCAGTATCTTCTCCTTAATGAAGATCTAAAACATGATAAAGATTTTTTTATT
CAAAACAAACACACAAATTATGAGGAATCTGATGAATACATTGATAATAACATGCTTGAG
TTTGGGAAAAAGTTGTATCAATTCAGTGCATAGACAACGCTTTAGTTTGCGCCATTTGTC
TACGTCTTAATTTTTTCCACCTTTTCAAACTCAAACTTTTCATTTTTTCCACCTTTTCAA
ACTCAAACTTTTCATTTTTTCCACCTTTTCAAACTCAAACTTTTCATTTTTTCCACCTTT
TCAAACTCAAACTTTTCATTTTTTCCACCTTTTCAAACTCAAACTTTTCATTTTTTCCAC
CTTTTCAAACTCAAACTTTTCATTTTTTCCACCTTTTCAAACTCAAACTTTTCATTTTTT
CCACCTTTTCAAAATCAAACTTTTCATTTTTTCCACTTTTTCAAGCTTTTTCAAACTTCA
TTTTTTTCAAACCTCGCCAGCGCGTCGCGCCCAACTAAGTAAACAGTACTCACCCCAAAC
GTATTTACTTTAAATACGCGCATGATATCCCTCTAAGTAGCATTTTATTATTTGAAACGG
TTTGGTATATACACACCAGGTACCCCGAAGGGCGAATTCTGCAGATATCCATCACACTGG
CGGCCGCCACCGCGGTGGAGCTCCAATTCGCCCTATAGTGAGTCGTATTACAATTCACTG
GCCGTCGTTTTACAACGTCGTGACTGGGAAAACCCTGGCGTTACCCAACTTAATCGCCTT
GCAGCACATCCCCCTTTCGCCAGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCT
TCCCAACAGTTGCGCAGCCTGAATGGCGAATGGGACGCGCCCTGTAGCGGCGCATTAAGC
GCGGCGGGTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCC
GCTCCTTTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCT
CTAAATCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAA
AAACTTGATTAGGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGC
CCTTTGACGTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACA
CTCAACCCTATCTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTAT
TGGTTAAAAAATGAGCTGATTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACG
CTTACAATTTAGGTGGCACTTTTCGGGGAAATGTGCGCGGAACCCCTATTTGTTTATTTT
TCTAAATACATTCAAATATGTATCCGCTCATGAGACAATAACCCTGATAAATGCTTCAAT
AATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCCGTGTCGCCCTTATTCCCTTTT
TTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAGTAAAAGATG
CTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAAGA
TCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCACTTTTAAAGTTCTGC
TATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTCGCCGCATAC
ACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTACGGATG
GCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACACTGCGGCCA
ACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACATGG
GGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACG
ACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATTAACTG
GCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAG
TTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTG
GAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCCT
CCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGAC
AGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACT
CATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGA
TCCTTTTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGT
CAGACCCCGTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCT
GCTGCTTGCAAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGC
TACCAACTCTTTTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTCC
TTCTAGTGTAGCCGTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACC
TCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCG
GGTTGGACTCAAGACGATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTT
CGTGCACACAGCCCAGCTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTG
AGCTATGAGAAAGCGCCACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCG
GCAGGGTCGGAACAGGAGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTT
ATAGTCCTGTCGGGTTTCGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAG
GGGGGCGGAGCCTATGGAAAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTT
GCTGGCCTTTTGCTCACATGTTCTTTCCTGCGTTATCCCCTGATTCTGTGGATAACCGTA
TTACCGCCTTTGAGTGAGCTGATACCGCTCGCCGCAGCCGAACGACCGAGCGCAGCGAGT
CAGTGAGCGAGGAAGCGGAAGAGCGCCCAATACGCAAACCGCCTCTCCCCGCGCGTTGGC
CGATTCATTAATGCAGCTGGCACGACAGGTTTCCCGACTGGAAAGCGGGCAGTGAGCGCA
ACGCAATTAATGTGAGTTAGCTCACTCATTAGGCACCCCAGGCTTTACACTTTATGCTTC
CGGCTCGTATGTTGTGTGGAATTGTGAGCGGATAACAATTTCACACAGGAAACAGCTATG
ACCATGATTACGCCAAGCTCGAAATTAACCCTCACTAAAGGGAACAAAAGCTGGTACCCG
TACATGTACGCATATATTATACAAAATTAAAAATATAAAATGCACAATATAATGTATGTA
TTTAAAATTATATATATATATGAATACAACATTAAAAAGACGCTTATGAAAAGTTTATGA
ATATATAAAATTTTTATCTTTACTTTTATTTATATATATATCAAGCTGAAATTAAAGAGA
GTGTCTTTTTTTCATCTTATATATAGTGAGATATATTTTTTTGTTTTCTTTTTTTGTTTC
TCTTTTTCATTTTTGTTTAACGGTTTTCTGTAAATGCACTTGTATATCCGTTGTTCAATT
TGAAGGTATTGTTAACCTTTAAAAAAAAAAGAAAAAAAGAAAAAAATATATTACAGGTCA
GAAAAAAGGCATATTCTTATTTGGGATTAAAACATTTAATAAATTTCAAAAAAAAATTAG
AGGCTTCTAAACTCAAAATCAACACATTCTTTTAATTATTTGAATTGTATTATTAAATTA
GGAAATCATATATATATGCAAAAATAGTAAAAAAACAAACAAATAAATAAATAAATAAAT
AAATAAATAAATAAATAAATAAATAAATAAATAAATAAATAAATAAATAAATAAATAAAT
AAATAAATAAGAGCATATATTATGGGTTTTCTAAAAATTGGAACTCCATTAAGTTGGGAA
GATGTGCAGAATGTCAAATCATTAATTAGATTATATGGAATATTGCAATTTGTTCATTCA
TATAAATGTAATAAGGATAGAAAAGATGAACATATTATGTTTGGAGATGAAATAGAATAT
ATAATTATAAGAAATGATGATAAATTAAAAGAAGCATCTGCATTATTATGTGCAACTGAT
TTAATAGATGAAATGATGAATTTAGAAAGTGGAAATGATTGCCA
Restriction sites to linearize plasmid KpnI/SacII
Partial or complete disruption of the geneComplete
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 1GGGGTACCCGTACATGTACGCATATATTATACA
Additional information primer 12751 (KpnI); 5'
Sequence Primer 2CCCAAGCTTGGCAATCATTTCCACTTTCTAAATTCATC
Additional information primer 22752 (HindIII); 5'
Sequence Primer 3CATGCCATGGATGGGTTTTCTAAAAATTGGAACTCC
Additional information primer 32562 (KpnI); 3'
Sequence Primer 4CGGGGTACCTGGTGTGTATATACCAAACCGTTTC
Additional information primer 42563 (KpnI); 3'
Sequence Primer 5
Additional information primer 5
Sequence Primer 6
Additional information primer 6