By Curtis A. Machida
Oregon overall healthiness & technology Univ., Portland, OR. textual content presents uncomplicated instruments wanted for layout unique gene supply vectors, and an knowing of ways to use viral vectors to the remedy of genetic issues. comprises step by step equipment, vectors from significant different types, assistance at the choice, and examples. DNLM: Genetic Vectors.
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Extra info for Viral Vectors for Gene Therapy: Methods and Protocols (Methods in Molecular Medicine)
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1993) An HSV LAT null mutant reactivates slowly from latent infection and makes small plaques on CV-1 monolayers. Virology 192, 618–630. 57. Drolet, B. , Perng, G. , Villosis, R. , Slanina, S. , Nesburn, A. , and Wechsler, S. L. (1999) Expression of the first 811 nucleotides of the herpes simplex virus type 1 latency-associated transcript (LAT) partially restores wild-type spontaneous reactivation to a LAT-null mutant. Virology 253, 96–106. 58. Loutsch, J. , Perng, G. , Hill, J. , Marquart, M. , Block, T.
This cascade is well ordered and tightly regulated and dictates the controlled expression of three classes of genes; the IE or α genes, the early (E) or β genes, and the late (L) or γ genes (2). There are five IE genes; infected cell polypeptides (ICP) 0, 4, 22, 27, and 47. Four of these IE genes (ICP0, 4, 22, and 27) are involved in the transcriptional and posttranscriptional regulation of the expression of the E and L genes. The remaining IE gene, ICP47, is not a regulatory IE protein, but inhibits the transporters of antigen processing (TAP), thereby helping the virus avoid the host’s immune surveillance mechanisms (3).
If necessary, pure viral DNA can be prepared from virus harvested from the media of infected cells or from virus particles that have been gradient purified. The yield of DNA obtained in this instance is significantly reduced. 2. As the viral DNA is 152 kb in size, the use of wide-bore pipet tips (Bio Rad, Hercules, CA) is recommended to prevent shearing of the viral DNA, thereby increasing the likelihood of delivering intact genomes to cells during transfection. 3. In our experience, spooling the DNA gives a higher quality yield with improved transfection efficiency.