Ribozyme vectors

Li MJ, Kim J, Li S, Zaia J, Yee JK, Anderson J, Akkina R, Rossi JJ (2005b) Long-term inhibition of HlV-1 infection in primary hematopoietic cells by lentiviral vector delivery of a triple combination of anti-HIV shRNA, anti-CCR5 ribozyme, and a nucleolar-localizing TAR decoy. Mol Ther 12 900-909... 

In most cases, the DNA inserted into the MCS for expression is not a genomic gene with the original exon/intron configuration, but a cDNA. In exceptional cases, antisense RNA or ribozymes may be expressed. cDNA lacks intronic sequences, but still has the 5 and 3 UTR sequences. As discussed above, the presence of the 3 UTR may reduce the stability of mRNA transcribed from the cDNA insert. Furthermore, the length and secondary structure of the 5 UTR may influence the efficiency of translation. Therefore, it is generally recommended to use cDNA with only short 5 and 3 UTR sequences for expression with expression vectors. 

Stable intracellular expression of transcriptionally active ribozymes can be achieved by viral vector-mediated delivery. Currently, retroviral vectors are the most commonly used in cell culture, primary cells... 

Gibson, S.A., Pellenz, C., Hutchison, R.E., Davey, F.R. and Shillitoe, E.J. (2000) Induction of apoptosis in oral cancer cells by an anti-bcl-2 ribozyme delivered by an adenovirus vector. Clin. Cancer Res., 6, 213-222. 

RNA pol III promoters, including tRNA, adenovirus VAl, Ul, and U6 small nuclear RNA derivatives, have allowed ubiquitous and very high intracellular levels of ribozyme expression. Several creative permutations, such as the combination of two promoters driving the same ribozyme gene or the placement of the ribozyme expression cassette within the U3 region of a Long terminal repeat, to produce a double-copy retroviral vector led to further increases in ribozyme expression. Finally, multiple ribozymes can be expressed within the same RNA transcript or from multiple promoter-ribozyme cassettes. 

Another state-of-the-art technique introduced in in vitro (and in vivo as well) transcriptions is the use of an RNA transcript-trimming plasmid which incorporates a ribozyme technology (70,71). A typical vector construct has the configura-... 

Zhang Y, Nemunaitis J, Tong A. Generation of a ribozyme-adenoviral vector against K-ras mutant human lung cancer cells. Mol Biotechnol 2000 15 39-49. 

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