Synthetic Gene DataBase
 

Synthetic Gene 49


 
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Field NameNatural GeneSynthetic Gene
SGDB Gene ID4149
GenBank AccessionAJ277646AF512507
GenBank GI993118425271700
Gene NameAMA1 (FVO)PpAMA1 (FVO)
Gene Length (bp)18161566
SpeciesPlasmodium falciparum (malaria)Pichia pastoris
StrainsFVOGS114 (his4)
CDSattattgagcgcctttgagtttacatatatgataaactttggaagaggacagaattattg
ggaacatccatatcaaaaaagtgatgtgtatcatccaatcaacgaacatagggaacatcc
aaaagaatacgaatatccattacaccaggaacatacataccaacaagaagattcaggaga
agacgaaaatacattacaacacgcatatccaatagaccacgaaggtgctgaacccgcacc
acaagaacaaaatttattttcaagcattgaaatagtagaaagaagtaattatatgggtaa
tccatggacggaatatatggcaaaatatgatattgaagaagttcatggttcaggtataag
agtagatttaggagaagatgctgaagtagctggaactcaatatagacttccatcagggaa
atgtccagtatttggtaaaggtataattattgagaattcaaatactacttttttaaaacc
ggtagctacgggaaatcaagatttaaaagatggaggttttgcttttcctccaacaaatcc
tcttatatcaccaatgacattaaatggtatgagagatttttataaaaataatgaatatgt
aaaaaatttagatgaattgactttatgttcaagacatgcaggaaatatgaatccagataa
tgataaaaattcaaattataaatatccagctgtttatgattacaatgataaaaagtgtca
tatattatatattgcagctcaagaaaataatggtcctagatattgtaataaagaccaaag
taaaagaaacagcatgttttgttttagaccagcaaaagataaattatttgaaaactatac
atatttaagtaaaaatgtagttgataactgggaagaagtttgccctagaaagaatttaga
gaatgcaaaattcggattatgggtcgatggaaattgtgaagatataccacatgtaaatga
attttcagcaaatgatctttttgaatgtaataaattagtttttgaattgagtgcttcgga
tcaacctaaacaatatgaacaacatttaacagattatgaaaaaattaaagaaggtttcaa
aaataagaacgctagtatgatcaaaagtgcttttcttcccactggtgcttttaaagcaga
tagatataaaagtcatggtaagggttataattggggaaattataacagagaaacacaaaa
atgtgaaatttttaatgtcaaaccaacatgtttaattaacaattcatcatacattgctac
tactgctttgtcccatcccatcgaagttgaacacaattttccatgttcattatataaaga
tgaaataaagaaagaaatcgaaagagaatcaaaacgaattaaattaaatgataatgatga
tgaagggaataaaaaaattatagctccaagaatttttatttcagatgataaagacagttt
aaaatgcccatgtgaccctgaaatggtaagtaatagtacatgtcgtttctttgtatgtaa
atgtgtagaaagaagggcagaagtaacatcaaataatgaagttgtagttaaagaagaata
taaagatgaatatgcagatattcctgaacataaaccaacttatgataacatgaaaattat
aattgcatcatcagctgctgtcgctgtattagcaactattttaatggtttatctttataa
aagaaaaggaaatgctgaaaaatatgataaaatggatcaaccacaacattatgggaaatc
aacatcaagaaatgatgaaatgttagatcctgaggcatctttttggggggaagaaaaaag
agcatcacatacaaca
cagaactactgggaacatccataccaaaagtctgatgtgtaccatccaataaatgaacat
cgtgaacatccaaaggaatatgaatacccattgcaccaggaacatacttatcaacaagaa
gattccggggaagacgaaaacactttgcaacacgcatacccaattgaccacgaaggagcg
gaacccgcaccacaagaacaaaacttgttttccagcatcgaaattgtcgaaagatctaac
tacatgggaaacccatggacggaatacatggcaaagtacgatatcgaagaagttcatggt
tccggaattagagtcgatttgggggaagatgcggaagtcgcggggacacaatacagacta
ccatccggtaagtgtccagtctttggaaagggaattatcatcgagaactccaagacaaca
tttttgactccggtcgcgacggggaaccaagatttgaaggatggcggatttgcgtttcct
ccaactaaccctcttatttccccaatgactttgaacggaatgagagatttttacaagaac
aacgaatacgtcaagaacttggatgaactgacattgtgttccagacatgcagggaacatg
aacccagataacgataagaactccaactacaagtacccagcggtttacgattataacgat
aagaaatgtcatattttgtacatcgcagcgcaagaaaacaacggacctagatactgtaac
aaggaccaatctaagagaaatagcatgttttgttttagaccagcaaaggataagttgttt
gaaaatttggtctacttgtctaagaacgtcgttgataattgggaagaagtttgccctaga
aagaacttggagaacgcaaagttcgggttgtgggtagatgggaactgtgaagatattcca
catgtcaacgaattttccgcaaacgatctttttgaatgtaacaagttggtttttgaactg
tctgcgtcggatcaacctaagcaatacgaacaacatttgactgattacgaaaagatcaag
gaaggattcaagaacaaaaatagagaaatgataaagtctgcgtttcttcccacaggagcg
tttaaggcagatagatacaagtctcatggaaaaggatacaactgggggaactacaataga
gaaactcaaaagtgtgaaatctttaacgtaaagccaacttgtttgatcaatgacaagaac
tatatcgcgacaacagcgctgagtcatcccatagaagttgaacacaactttccatgttcc
ttgtacaaggatgaaattaaaaaggaaatagaaagagaatccaagcgaatcaagttgaac
gataacgatgatgaaggtaacaagaagatcattgcgccaagaatctttatctccgatgat
aaggactctttgaagtgcccatgtgaccctgaaatggtctctcaatctacttgtaggttc
tttgtctgtaagtgtgtcgaaagacgtgcagaagtcacttccaacaacgaagttgtcgtt
aaggaagaatacaaggatgaatacgcagatatccctgaacataagccaacatacgataat
atgaag
5' End
3' End
NotesThe CDS was incomplete. But only AA25 to AA546 was recoded for expression.Only the AA25 to AA546 were used in synthetic gene. The synthetic gene contains YV at the N-terminus and TSHHHHHH at the C-terminus.
Expression VectorNApPIC9K
Assay MethodsNASDS-PAGE, Western blot
ResultsNot determinedProduction of final AMA1 protein is 50 mg/liter.
Protein FunctionHelp invasion (apical membrane antigen)
Recoding PurposeTo improve expression
Synthesized ByAuthors
Recoding MethodThe synthetic DNA sequences were designed using a computer algorithm that matched each codon used by
P. falciparum with a codon used with the same frequency for that amino acid by P. pastoris.
Publication Author(s)Kennedy MC, Wang J, Zhang Y, Miles AP, Chitsaz F, Saul A, Long CA, Miller LH, Stowers AW.
Corresponding AuthorAnthony W. Stowers
Corresponding AddressMalaria Vaccine Development Unit, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Rockville, Maryland 20852, USA.
Publication Year2002
Publication TitleIn vitro studies with recombinant Plasmodium falciparum apical membrane antigen 1 (AMA1): production and activity of an AMA1 vaccine and generation of a multiallelic response.
AbstractApical membrane antigen 1 (AMA1) is regarded as a leading malaria blood-stage vaccine candidate. While the overall structure of AMA1 is conserved in Plasmodium spp., numerous AMA1 allelic variants of P. falciparum have been described. The effect of AMA1 allelic diversity on the ability of a recombinant AMA1 vaccine to protect against human infection by different P. falciparum strains is unknown. We characterize two allelic forms of AMA1 that were both produced in Pichia pastoris at a sufficient economy of scale to be usable for clinical vaccine studies. Both proteins were used to immunize rabbits, singly and in combination, in order to evaluate their immunogenicity and the ability of elicited antibodies to block the growth of different P. falciparum clones. Both antigens, when used alone, elicited high homologous anti-AMA1 titers, with reduced strain cross-reactivity. Similarly, sera from rabbits immunized with a single antigen were capable of blocking the growth of homologous parasite strains at levels theoretically sufficient to clear parasite infections. However, heterologous inhibition was significantly reduced, providing experimental evidence that AMA1 allelic diversity is a result of immune pressure. Encouragingly, rabbits immunized with a combination of both antigens exhibited titers and levels of parasite inhibition as good as those of the single-antigen-immunized rabbits for each of the homologous parasite lines, and consequently exhibited a broadening of allelic diversity coverage.
JournalInfect Immun.. 70(12): 6948-60.
SummaryTo achieve high production of AMA1 protein, a profile matching algorithm was used to modify the native AMA1 gene with the reference to P. pastoris codon preference. it appeared that synthetic AMA1 genes were expressed successfully in P. pastoris.
Comments
Discussion http://www.evolvingcode.net/forum/viewtopic.php?t=502
PubMed ID12438374
Submitter NameWu, Gang
Submitter AddressDepartment of Biological Sciences, University of Maryland Baltimore County, 1000 Hilltop Circle, Baltimore, MD 21250 USA
Entry ConfirmationNo
 
 

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