Summary

RMgm-110
Malaria parasiteP. berghei
Genotype
DisruptedGene model (rodent): PBANKA_0712200; Gene model (P.falciparum): PF3D7_0818600; Gene product: BEM46-like protein, putative
Phenotype Oocyst; Sporozoite; Liver stage;
Last modified: 18 February 2009, 12:04
  *RMgm-110
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) : 18974882
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/ResearcherE. Lasonder, C.J. Janse, H.G. Stunnenberg
Name Group/DepartmentDepartment of Molecular Biology
Name InstituteNCMLS, Radboud University Nijmegen
CityNijmegen
CountryThe Netherlands
Name of the mutant parasite
RMgm numberRMgm-110
Principal name808; 843cl1
Alternative name
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
OocystOocyst numbers in A. stephensi were comparable to wild type (mean of 188 oocysts per mosquito compared to 169 in wild type). Sporozoite formation is blocked inside the oocysts and only few oocysts were found that contained sporozoites. Strongly reduced numbers of hemocoel and salivary gland sporozoites (see phenotype 'Sporozoite').
SporozoiteSporozoite formation is blocked inside the oocysts and only few oocysts were found that contained sporozoites. Strongly reduced numbers of hemocoel and salivary gland sporozoites in infected A. stephensi mosquitoes (mean of 2625 sporozoites per salivary gland compared to over 10.000 in wild type).
Liver stageStrongly reduced numbers of hemocoel and salivary gland sporozoites in infected A. stephensi mosquitoes (mean of 2625 sporozoites per salivary gland compared to over 10.000 in wild type). No blood stage infections developed when mice were infected by bite of infected mosquitoes. Also intravenous injection of sporozoites (1–2x10(4)) did not result in blood stage infections. Sporozoites showed in vitro hepatocyte HepG2) traversal and invasion.
Additional remarks phenotype

Mutant/mutation
The mutant lacks expression of PB402680.00.0

Protein (function)
Unknown. The protein was detected in a proteome analysis of P. falciparum oocysts and sporozoites and was present in oocyst-derived sporozoites and in salivary gland sporozoites. It contains a signal peptide and a transmembrane region. 

Phenotype
The phenotype analyses indicate a role of the protein in the formation of sporozoites within the oocysts. The mutant produces normal numbers of oocysts; however,  very few oocysts  containing sporozoites are present and only few free sporozoites are found in the hemocoel and in the salivary glands. The few sporozoites formed are not infectious to mice. However, sporozoites were able to traverse and invade hepatocytes in vitro. This suggests that the lack of sporozoite infectivity to mice may be due to an additional defect in liver stage development after invasion of the hepatocyte.


  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_0712200
Gene Model P. falciparum ortholog PF3D7_0818600
Gene productBEM46-like protein, putative
Gene product: Alternative name
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
AATGATGTTTTTTCCTTCAATTTCGATATCGTTCACTCTAGTTTCACTTTAGTGCATATT
TTGTATCCCTTTCCGTGTGCTTTCCTTTTTCATCTTTATGTTTTTTTTTCTTCGTTTTTA
AATTTGTTAGATATGCCCAAGCAAAAAGTTTATTTATACTGCAAAAGACGGAGAGCATAA
TGACATTCTTGTAAATGATGATGGATCATATCATCGATCAATGAAAATATTTATTGAAAC
AGTCAACTCTACATATAGCAAAAAATAATTATAATTCAAATGAAGGAAATTAATTATGTC
AATCTTTGTGTGTGTGTGTGCAATCGCCTATTTATGTAAAAAATATATATTTTAAGATAG
AAATTTTGCAAAGACATTTTTTATTTATCTTATATTGTTTATTAAAGTTTATCCAATTAA
TTCAACGTTTGCAATTATTTTCCTAAAAAATAAAGAAATCCCAAATTATACACATATTGA
AGAAAATTCATTTTAAAACAATATTATTTTTTATAATTCAAATACCATACTTTTTATTTG
TCTACCTAAGTTGGATATATAAAAATATTGTATATGACAAATCTACAATAATATTAAAAA
AAAAAATATTATTATATTTTTATTCATAATTATTTTTGAAATTCAAGTTATAAACTGTTT
TGTATTAATAATAACCATTTGCTTTTCATAATTAATTTACTTTTATATTATTATGTCACT
TTTTATTTGTATTAACAATTAAAATACAAATTCCATTTTGTTGTTTTAATACCATTCTGT
ATTATGTTTTTATGCGAATACTTTATTCATAACAAATAATATATAAGCTAAACAATACAT
GAATTAATAACTTTTGAATTAAATATAAAGAATATAATACGAAAAAAAAAGTTTATATTA
TTTTAGCATTATATTGCATAACCATTATTCTTCTTCAAAAAGAGGGATAAATAAAATGGT
CTATTTTGGTAATACAACAAATTTATTTAACACGAAAATATATACCATTTATTATAAATT
ATTTATCGAAAAATTTGTAAATGTATATATGATTTGAACTACATAAAATATACTTTTAAA
ACTTCTTCGCTAGTTCAAGGCTATAACATGGATCCACTAGTTCTAGAGCGGCCGCCACCG
CGGTGGAGCTCCAATTCGCCCTATAGTGAGTCGTATTACGCGCGCTCACTGGCCGTCGTT
TTACAACGTCGTGACTGGGAAAACCCTGGCGTTACCCAACTTAATCGCCTTGCAGCACAT
CCCCCTTTCGCCAGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAG
TTGCGCAGCCTGAATGGCGAATGGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGT
GTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTC
GCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGG
GGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGAT
TAGGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACG
TTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCT
ATCTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAA
AATGAGCTGATTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATT
TAGGTGGCACTTTTCGGGGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATAC
ATTCAAATATGTATCCGCTCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAA
AAAGGAAGAGTATGAGTATTCAACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCAT
TTTGCCTTCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATC
AGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAAGATCCTTGAGA
GTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCACTTTTAAAGTTCTGCTATGTGGCG
CGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTCGCCGCATACACTATTCTC
AGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTACGGATGGCATGACAG
TAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACACTGCGGCCAACTTACTTC
TGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACATGGGGGATCATG
TAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGACGAGCGTG
ACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATTAACTGGCGAACTAC
TTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGTTGCAGGAC
CACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCGGTG
AGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCCTCCCGTATCG
TAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACAGATCGCTG
AGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATATATAC
TTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTG
ATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCG
TAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGC
AAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTC
TTTTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTCCTTCTAGTGT
AGCCGTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGC
TAATCCTGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACT
CAAGACGATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACAC
AGCCCAGCTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAG
AAAGCGCCACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCG
GAACAGGAGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTG
TCGGGTTTCGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGA
GCCTATGGAAAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTT
TTGCTCACATGTTCTTTCCTGCGTTATCCCCTGATTCTGTGGATAACCGTATTACCGCCT
TTGAGTGAGCTGATACCGCTCGCCGCAGCCGAACGACCGAGCGCAGCGAGTCAGTGAGCG
AGGAAGCGGAAGAGCGCCCAATACGCAAACCGCCTCTCCCCGCGCGTTGGCCGATTCATT
AATGCAGCTGGCACGACAGGTTTCCCGACTGGAAAGCGGGCAGTGAGCGCAACGCAATTA
ATGTGAGTTAGCTCACTCATTAGGCACCCCAGGCTTTACACTTTATGCTTCCGGCTCGTA
TGTTGTGTGGAATTGTGAGCGGATAACAATTTCACACAGGAAACAGCTATGACCATGATT
ACGCCAAGCTCGAAATTAACCCTCACTAAAGGGAACAAAAGCTGGTACCCATCATAGTTG
CTGTCATAGCATCCATAGCTTTGATAAACAGTAAGAACATATATAACATTTTAAGAAAAT
TAAATTCAAATACATAATTATTAATAAGCATTATGAATGTCTATATCGATAGTTAAATGT
GGTATACATTTTTTAATATTTTTTATTTTAAGCATATATATACTTTAGCCAGGATAATCT
TATTTTCGTAAAAAAAGGTAAATAATTATTAACATCTTATAAATATAAACTAAATTAATT
GAATACATTATGTATCAAACATTTTATCGACGAATATTATATGACACTAAATACAACCAT
GTATCATCTCCATTCGTATAATCGTTATATATGCATTTTCACTTTTTCATTTTGATTTTA
CTTGCTTTTCTGCTCTATTAGACATCGATCCCATATATAACAAACCCTTAGGTAACATAA
AAAAAAACAATTCTTAATAAAATTTGTAAACTTGTATTTTTAATTTACCAAACTATAATA
TGCATTATGTTATCCACAGTATGCTATATATATTATACTATGTTGCTCTTTTCATAATTT
AGGAGATAATTATGAAGAAGTTATGATAAAAACCGAAGATGGACATCAAAATAAATGGTA
GCCAATAAAAAAAAATCGGATTAAATTATATAGTAAAATTATTATTTTTATTTTAACTAC
TATTATATCTTCAATATTATGTTCCTTTTTTATTTTATCCATTAATAATTAGTTGGTTCA
TAAAAGCAAAAGATTATGAAAGCAAACCAGTTATTCTTTATTTTCTTGGAAATGGAAGTT
GTAAGAATTAAAAAATTTTATTCAAAATAAATTTTTATATATGTGGATAAAAAAAGTATG
ATAAGGCTGAGTTTATTATACGAATTTCCCTACAATATATATATAACCCAATTGATGATT
TTCTATCTCATTTTTTTTTTTTTTTTTTAATATAAATATATTTTTATAGACGTAGAAAAG
AATGTTAATATATTCAATTTGATGGTTGGAAGAGTAGATGTGAGCATA
Restriction sites to linearize plasmid
Partial or complete disruption of the geneComplete
Additional remarks partial/complete disruption See for more information about the disruption Lasonder, E. et al., 2008, PloS Pathogens 10, e10000195 (Supporting Information: S3).
Selectable marker used to select the mutant parasitetgdhfr
Promoter of the selectable markerpbdhfr
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modificationSee for more information about the disruption Lasonder, E. et al., 2008, PloS Pathogens 10, e10000195 (Supporting Information: S3).
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 1CGGGGTACCCATCATAGTTGCTGTCATAGCATCC
Additional information primer 1L2632 (Asp718); 5'
Sequence Primer 2CCCAAGCTTATGCTCACATCTACTCTTCCAACC
Additional information primer 2L2633 (HindIII); 5'
Sequence Primer 3CCGGATATCGTTCACTCTAGTTTCACTTTAGTGC
Additional information primer 3L2634 (EcoRV); 3'
Sequence Primer 4CGCGGATCCATGTTATAGCCTTGAACTAGCG
Additional information primer 4L2635 (BamHI); 3'
Sequence Primer 5
Additional information primer 5
Sequence Primer 6
Additional information primer 6