Please use this identifier to cite or link to this item: http://hdl.handle.net/10603/2560
Title: Studies on inhibition of gene expression by DNAzymes
Researcher: Chaudhury, Indrajit
Guide(s): Hari Das, Rakha
Keywords: DNAzymes
Biology
Genomics
Upload Date: 1-Sep-2011
University: University of Pune
Completed Date: 2008
Abstract: DNAzymes are single‐stranded 2‐deoxyoligonucleotides with catalytic activity. 10‐23 DNAzyme being the most efficient RNA‐phosphodiesterase has been extensively utilized in gene silencing. This study demonstrates the effectiveness of 10‐23 DNAzyme in silencing several genes of therapeutic interest like inducible nitric oxide synthase (iNOS), tumor necrosis factor‐α (TNF‐α) and its receptors TNF‐R1 and TNF‐R2, hepatitis C virus (HCV) RNA. The designed DNAzymes Dz I, II and III cleaved murine iNOS mRNA both in vitro and in vivo. DNAzyme target site I or translation initiation site and site II have computer predicted (MFOLD) secondary structures but site III has no secondary structure. All the three DNAzymes cleaved the short transcripts generated from cloned DNA almost with equal efficiency while cleavage efficiency is higher at site III than the other two sites on isolated iNOS mRNA. Interestingly, at intracellular level, DNAzyme targeted at translation initiation codon (site I) having secondary structure cleaved iNOS mRNA, and suppressed its enzymatic activity and protein expression more efficiently than that targeted at sites II and III. Due to the efficient cleavage activity of the DNAzymes as observed in murine macrophage J774 cells, the DzI and DzIII that target site I and III were further tested in in vivo LPS induced BALB/c mice. Consistent with the previous results in J774 cells, DzI suppressed iNOS expression much more efficiently than DzIII. TNF‐α is a pro‐inflammatory cytokine secreted primarily by activated macrophages. Signaling through the TNF‐α receptors, TNF‐R1 and TNF‐R2 have been implicated in various acute and chronic inflammatory disorders. TNFreceptor mediated activation of NF‐kB is the key step that results in the transcriptional activation of a number of genes which contribute to the succession of inflammatory cascade. We have designed DNAzymes targeting mRNAs of TNF‐α and its receptors.
Pagination: 91p.
URI: http://hdl.handle.net/10603/2560
Appears in Departments:Institute of Genomics and Integrative Biology

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01_title.pdfAttached File153.48 kBAdobe PDFView/Open
02_certificate.pdf129.48 kBAdobe PDFView/Open
03_declaration.pdf130.3 kBAdobe PDFView/Open
04_dedication.pdf70.92 kBAdobe PDFView/Open
05_contents.pdf127.58 kBAdobe PDFView/Open
06_acknowledgement.pdf135.59 kBAdobe PDFView/Open
07_abstract.pdf178.27 kBAdobe PDFView/Open
08_preface.pdf176.57 kBAdobe PDFView/Open
09_abbreviation.pdf182.25 kBAdobe PDFView/Open
10_list of figures and tables.pdf180.85 kBAdobe PDFView/Open
11_chapter 1.pdf383.13 kBAdobe PDFView/Open
12_chapter 2.pdf425.37 kBAdobe PDFView/Open
13_chapter 3.pdf741.29 kBAdobe PDFView/Open
14_chapter 4.pdf750.7 kBAdobe PDFView/Open
15_chapter 5.pdf501.17 kBAdobe PDFView/Open
16_chapter 6.pdf198.14 kBAdobe PDFView/Open
17_references.pdf269.46 kBAdobe PDFView/Open
18_appendix.pdf192.84 kBAdobe PDFView/Open


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