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Download Antisense and Ribozyme Methodology: Laboratory Companion by Ian Gibson PDF

By Ian Gibson

Antisense and ribozymes have a comparatively brief but profitable background as study instruments in gene expression reports, and therefore are regarded as having excessive strength reagents in treating viral infections and melanoma. This laboratory spouse offers precise details at the power, benefits and barriers of this technique. It seriously discusses capability pitfalls, offers innovations for selecting ambitions and supply structures, so that it will enable the choice of the optimal technique for reaching quickly and trustworthy experimental good fortune with any human or different organic process. For researchers, technicians and complex graduates in experimental medication, molecular and cellphone biology.

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1 Choosing Antisense Arms of Hammerhead Ribozymes The two antisense arms of a hammerhead ribozyme should be sufficiently long to form a stable ribozyme-substrate complex via approximately 15 or more Watson-Crick base pairs. However, there are no general rules on the exact length and symmetry of both antisense arms. Biologically successful examples of the use of ribozymes as inhibitors of gene expression and viral functions have been described for all kinds of combinations. Usually, the selection of antisense arms is influenced by technical considerations that include the location of the cleavable motif or the location of restriction sites.

1996 and unpublished). The selection procedure included ribozymes directed against all potential cleavage sites located on the target molecule. Both steps of ribozyme action, substrate binding and the cleavage step, were included in the selection. , 7 to 8 nt on either side. 1 Choosing Antisense Arms of Hammerhead Ribozymes The two antisense arms of a hammerhead ribozyme should be sufficiently long to form a stable ribozyme-substrate complex via approximately 15 or more Watson-Crick base pairs.

1995). Two additional experimental options are available to support the construction of recombinant genes for long-chain ribozymes. Firstly, cDNA fragments can be converted into ribozymes if they contain specific recognition sequences for restriction endonucleases. This strategy makes use of the fact that the recognition sequences for a series of restriction endonucleases contain a cleavable triplet for hammerhead ribozymes (Tabler and Tsagris, 1991). For example, the recognition sequence for Sal I (GTCGAC) contains the triplet GTC, which is GUC at the RNA level, and which represents a potential cleavable motif.

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