Summary

RMgm-584
Malaria parasiteP. berghei
Genotype
DisruptedGene model (rodent): PBANKA_0606700; Gene model (P.falciparum): PF3D7_1208200; Gene product: cysteine repeat modular protein 3 (CRMP3)
Phenotype Oocyst; Sporozoite; Liver stage;
Last modified: 3 April 2011, 14:07
  *RMgm-584
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) : 21453484
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/ResearcherJ. Thompson; K.D. Augustijn
Name Group/DepartmentInstitute of Immunology and Infection Research
Name InstituteSchool of Biological Sciences, University of Edinburgh
CityEdinburgh
CountryUK
Name of the mutant parasite
RMgm numberRMgm-584
Principal name324cl2
Alternative nameΔpcrmp3
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Phenotype
Asexual blood stageNot different from wild type
Gametocyte/GameteNot different from wild type
Fertilization and ookineteNot different from wild type
OocystNormal numbers of oocysts are produced. Inside oocysts morphologically normal sporozoites are formed. Sporozoites fail to egress from oocysts. No sporozoites are found in the salivary glands.
SporozoiteNormal numbers of oocysts are produced. Inside oocysts morphologically normal sporozoites are formed. Sporozoites fail to egress from oocysts. No sporozoites are found in the salivary glands.
Mechanically liberated sporozoites from oocysts show gliding motility and invade and infect hepatocytes but do not undergo further development.
Liver stageMechanically liberated sporozoites from oocysts show gliding motility and invade and infect hepatocytes in vitro but do not undergo further development.
Additional remarks phenotype

Mutant/mutation
The mutant lacks expression of CRMP3 (cysteine repeat modular protein 3)

Protein (function)
The Cysteine Repeat Modular Proteins (CRMP1–4) of Plasmodium, are encoded by a small gene family that is conserved in malaria and other Apicomplexan parasites. They are very large, predicted surface proteins with multipass transmembrane domains containing motifs that are conserved within families of cysteine-rich, predicted surface proteins in a range of unicellular eukaryotes, and a unique combination of protein-binding motifs, including a > 100 kDa cysteine-rich modular region, an epidermal growth factor-like domain and a Kringle domain.

Phenotype
The phenotype analyses indicate that CRMP3 is not essential for blood stage development and for the formation of oocysts. The phenotype analyses indicate that CRMP3 plays a role in egress of sporozoites from oocysts and also during development of the early liver stages.

Additional information
The phenotype of P. berghei mutants lacking expression of CRMP3 and CRMP4 (RMgm-585) is different from P. berghei mutants lacking expression of CRMP1 (RMgm-198) and CRMP2 (RMgm-199). Crmp1 and crmp2 gene knockout parasites  establish normal blood-stage infection and transmit to the mosquito to form oocysts that release large numbers of sporozoites into the mosquito haemocoel. These sporozoites are infectious to the mouse when injected intravenously but are unable to target and invade mosquito salivary glands and thus do not transmit via infected mosquito bite.

crmp4 was also disrupted in the GFP-reporter line P. berghei ANKA 507cl1 (RMgm-7) using the same method/plasmid as described here (line 658m1cl1). In addition, 'double knock' mutants have been generated lacking both CRMP3 and CRMP4 (line  656m2cl1, 656m2cl2). Details of these lines have not been published. CRMP3 has been tagged with GFP (line 436) and cmyc (line 1048) but parasites of these lines have not been analysed in detail.

Other mutants
RMgm-198: A mutant lacking expression of CRMP1
RMgm-199: A mutant lacking expression of CRMP2.
RMgm-585: mutant lacking expression of CRMP4


  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_0606700
Gene Model P. falciparum ortholog PF3D7_1208200
Gene productcysteine repeat modular protein 3
Gene product: Alternative nameCRMP3
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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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
AATGATGTTTTTTCCTTCAATTTCGATATCGAATTCCATTTTCCGTTCACACTTATGAGT
ATATTAGAACTTATTTGGTGTCAATCAACAAAATATAAAAACAAATTACCTATATTAATA
TTATATCACATGCCTTCACAAGTTTGTAGTTTTCAGAACAAATTATTTTTATCAGGAGTT
TTGTTTTCTGGCTTTTTTTTTTTTATTTATTTATTTTTATTTATAAAATATTTTTATGGC
ACATTTAAAAATTTTAAAGTTTTCCAATCCTACGATAGAGATTTTAAATGTTATTTTTTA
TTTAATGGATATAATTATCAGAATCGATGCTGGGATTTTGTTAATATAATAAAAATTGTA
TTGTTTGTTACATCATTTATGTGCCAATTTTACACAAATAAAATTAATAATTCAAAATAT
TTTATTTTTTCATGTATAATATTCATTATAATTACAGAAATTACACTTATTTTATTATAT
TCACCATATGACAAAAGATCTAATAATGTTTTACAAAAACTTAGTCTTTTGTCTATTTTT
TCAGTACTCATAACATACTTAACTACACACTTTAGCTTCTTTTTTGATTTTTACATAATA
AGTGCATTACCATTTATATTGTTTATTTATTTTCACATACATACTATAAATAAAATTGTC
CTAGAATTTGCATTATATAAAAATATTTTAATGAAACCTACAAGACAAAAACAAAAACTA
GATTCAGAAGAGTTTCTAAATGAAAATAGCATTTTATTTAATTCCTATTTAAATACAAAA
ACGAATATCCAAGATAATGATGAAGATATTATTTTCTCAAACAGTAAAAAAATAAGTAGA
AAATATAAAAGATCTAAAAAAGAGATTCATTCAAAAATAAAAGAGAATTATAATTTACTA
AATTTTGTTTTAAGAATTAATAAAATACCTTCATGTTCCATATTTTTTAATGAGTAAGGA
TCCACTAGTTCTAGAGCGGCCGCCACCGCGGTGGAGCTCCAATTCGCCCTATAGTGAGTC
GTATTACGCGCGCTCACTGGCCGTCGTTTTACAACGTCGTGACTGGGAAAACCCTGGCGT
TACCCAACTTAATCGCCTTGCAGCACATCCCCCTTTCGCCAGCTGGCGTAATAGCGAAGA
GGCCCGCACCGATCGCCCTTCCCAACAGTTGCGCAGCCTGAATGGCGAATGGGACGCGCC
CTGTAGCGGCGCATTAAGCGCGGCGGGTGTGGTGGTTACGCGCAGCGTGACCGCTACACT
TGCCAGCGCCCTAGCGCCCGCTCCTTTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGC
CGGCTTTCCCCGTCAAGCTCTAAATCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTT
ACGGCACCTCGACCCCAAAAAACTTGATTAGGGTGATGGTTCACGTAGTGGGCCATCGCC
CTGATAGACGGTTTTTCGCCCTTTGACGTTGGAGTCCACGTTCTTTAATAGTGGACTCTT
GTTCCAAACTGGAACAACACTCAACCCTATCTCGGTCTATTCTTTTGATTTATAAGGGAT
TTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTAACAAAAATTTAACGCGAA
TTTTAACAAAATATTAACGCTTACAATTTAGGTGGCACTTTTCGGGGAAATGTGCGCGGA
ACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGACAATAA
CCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCCGT
GTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACG
CTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTG
GATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATG
AGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAG
CAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACA
GAAAAGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATG
AGTGATAACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACC
GCTTTTTTGCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTG
AATGAAGCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACG
TTGCGCAAACTATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGAC
TGGATGGAGGCGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGG
TTTATTGCTGATAAATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTG
GGGCCAGATGGTAAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACT
ATGGATGAACGAAATAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAA
CTGTCAGACCAAGTTTACTCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTT
AAAAGGATCTAGGTGAAGATCCTTTTTGATAATCTCATGACCAAAATCCCTTAACGTGAG
TTTTCGTTCCACTGAGCGTCAGACCCCGTAGAAAAGATCAAAGGATCTTCTTGAGATCCT
TTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAAACCACCGCTACCAGCGGTGGTT
TGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCGAAGGTAACTGGCTTCAGCAGAGCG
CAGATACCAAATACTGTCCTTCTAGTGTAGCCGTAGTTAGGCCACCACTTCAAGAACTCT
GTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCCAGTGGC
GATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTTACCGGATAAGGCGCAGCGG
TCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGGAGCGAACGACCTACACCGAA
CTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCTTCCCGAAGGGAGAAAGGCG
GACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGCGCACGAGGGAGCTTCCAGGG
GGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCACCTCTGACTTGAGCGTCGA
TTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAACGCCAGCAACGCGGCCTTT
TTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGTTCTTTCCTGCGTTATCCCCT
GATTCTGTGGATAACCGTATTACCGCCTTTGAGTGAGCTGATACCGCTCGCCGCAGCCGA
ACGACCGAGCGCAGCGAGTCAGTGAGCGAGGAAGCGGAAGAGCGCCCAATACGCAAACCG
CCTCTCCCCGCGCGTTGGCCGATTCATTAATGCAGCTGGCACGACAGGTTTCCCGACTGG
AAAGCGGGCAGTGAGCGCAACGCAATTAATGTGAGTTAGCTCACTCATTAGGCACCCCAG
GCTTTACACTTTATGCTTCCGGCTCGTATGTTGTGTGGAATTGTGAGCGGATAACAATTT
CACACAGGAAACAGCTATGACCATGATTACGCCAAGCTCGAAATTAACCCTCACTAAAGG
GAACAAAAGCTGGTACCGTGATAGCGGCTTAGACCAGCCATATTCCTTATCACTATATTA
TTATGATGAAGCAAAAACAAACCTTTTATTTTTAAGTGAGTTATCTACACCAAGAATACT
AATTTTTGAAATAGATAATAATATTAAATTATATCAAGTATATAATGATTTACCATTTAA
TATTGCTACTAATGTTATTACAACATCCCGTTTTTTGGTAGTATGTGGAATTAATTTAAA
TAATGAAAGTATTACAACTTATATTACATTTATTGAAATACAAGATTTATCCGATATAAA
AATAAAATATAATGGCTTTCCTCGTTCATTCAAGTATAAAGAAGTAGTAAACATACAACC
TTTAATAAAAAGTGATAATATTATAAAATTTAAAATGAATCAATATGACCATAATATTCC
AGATAATAATATTATTACTGGATTGAAAATAAAAAAATATACTGGTATTATTTATGGGAC
AATAGAAATATCTGCTATATTTGATATAGAAATAACAGTATATAGCAATTTTAAAAAAAA
AATATTAATTTTAAAAAAAAACATTTCGTCATGCCCTAAAGGCCATTTTTATAGTAATAA
TAATAATTGTGAGTTATGTCCTATTGGATATTTTAGTAAATATAACAGGAAAGATGAATG
CTATAGCTGTGAAGATTATTTAAAAAACTCAATTACATTTTATCCTGGTTCTGATTCAGA
AAATGAATGTTTATGTAAACCTGGTTATTATTTAAGAGAAAATAAATGTGTAGAATGCCC
AGAAGGATTTTATAAAGATGTTATAGGAAACTTTAAGTGCCATTTGACATGTGAGCCCGT
GAAGAAAAGTACTGTAACTGGAGCCAAAAGTTATGA
Restriction sites to linearize plasmid KpnI, BamHI
Partial or complete disruption of the genePartial
Additional remarks partial/complete disruption CRM-3 cds: 10674bp
Disrupted region: 3553 - 8485
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 1CACCGAGGTACCGTGATAGCGGCTTAGACCAGCC
Additional information primer 1L1448 (KpnI); 5'target forward
Sequence Primer 2AGTTTAAAGCTTCATAACTTTTGGCTCCAGTTACAGTAC
Additional information primer 2L1449 (HindIII); 5'target reverse
Sequence Primer 3GTGTTTGAATTCCATTTTCCGTTCACACTTATGAGTATATTAG
Additional information primer 3L1450 (EcoRI); 3'target forward
Sequence Primer 4TTTCGGATCCTTCTCATTAAAAAATATGGAACATGAAGG
Additional information primer 4L1451 (BamHI); 3'target reverse
Sequence Primer 5
Additional information primer 5
Sequence Primer 6
Additional information primer 6