Characterisation and Functional Analysis of Isochorismate Synthase cDNA from Morinda citrifolia L.
Isochorismate synthase catalyzes the substrate chorismate in the presence of a metal ion (Mg2+) cofactor to isochorismate. Isochorismate is prominent in the shikimate pathway to produce siderophores, anthraquinones and salicylate. In previous study, a near complete cDNA encoding isochorismate synth...
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| Format: | Thesis |
| Language: | English |
| Published: |
2016
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| Subjects: | |
| Online Access: | http://ir.unimas.my/id/eprint/30586/1/Chai%20Suk%20Phin%20ft.pdf |
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| Summary: | Isochorismate synthase catalyzes the substrate chorismate in the presence of a metal ion (Mg2+) cofactor to isochorismate. Isochorismate is prominent in the shikimate pathway to produce siderophores, anthraquinones and salicylate. In previous study, a near complete
cDNA encoding isochorismate synthase (mcICS) has been isolated from ‘mengkudu’ or Morinda citrifolia. The cDNA was characterized and identified to be highest identity of 72% to Catharanthus roseus (AJ006065). An in silico analysis of mcICS cDNA was studied in this research to determine the 3D structure and active amino acid residues. The mcICS cDNA was further analyzed by cloning into pET-41a(+) vector for expression analysis in prokaryote system. pET-41a(+)/mcICS was constructed and yielded molecular weight approximately 55kDa. The mcICS cDNA also cloned into pGSA1131 vector and transformed into mungbean (Vigna radiata) through a direct cotyledons Agrobacterium tumefaciens- mediated transformation. This type of transformation was less laborious,efficient and it showed 3.33 % of transformation efficiency. The putative transgenic were grown to maturity to analyse the stability of transgene integration and expression of T1
generation. Results showed that some individulas from T1 generation as indicated by posttransciptional gene siliencing which observed as chimera as the transgenic mungbean construct from different cells of zygotes. Hence, these results suggests to analyze next T2
generation to provides more stability for transgenes integration and expression. |
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