Genes manipulation

Very little is known about plant terminators. It has not been determined whether an animal terminator can be functional in plants or not. Until such information is available, it is safer to use a plant terminator for the maximum expression of a foreign gene. The nopaline synthase nos terminator is most frequently used although several other plant terminators have also been isolated. 

There should be no internal translation initiation signal (ATG) between a promoter sequence and a structural gene since an internal ATG sequence significantly reduces translation efficiency. The distance between the control regions (promoter and terminator) and the structural gene should not be too long. Otherwise a produced m-RNA would be less stable. 

Many proteins synthesized in a cell are modified. For example, carbohydrates are often attached to the proteins found in cell membrane. Some proteins become phosphorylated. Such modifications either activate or inactivate the function of the protein. Therefore, a foreign gene should be properly manipulated to obtain a correct post-translational modification. For example, if an attachment of a carbohydrate to a protein is desired, one may design the process so as to secrete the protein out of a cell membrane. During this procedure the protein can be properly modified. However, such modification mechanisms can be different in each living organism. Therefore, careful selection of a proper cell line is necessary. 

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N. G. Carr and co-workers, eds., Principles of Gene Manipulation An Introduction to Genetic Engineering, University of California Press, Berkeley, Calif., 1980. 

Old R.W. Primrose S.B. (1989) Principles cf Gene Manipulation An Introduction to Genetic Engineering, 4th edn. Oxford Blackwell Scientific Publications. 

Iuchi, S., M. Kobayashi et al. (2001). Regulation of drought tolerance by gene manipulation of 9-cis-epoxycarotenoid dioxygenase, a key enzyme in abscisic acid biosynthesis in Arabidopsis. Plant J. 27(4) 325-333. 

Hutchinson, C.R. and Fujii, I. (1995) Polyketide synthase gene manipulation a structure-function approach in engineering novel antibiotics. Annual Review of Microbiology, 49, 201. 

Wendland, J. 2003. PCR-based methods facilitate targeted gene manipulations and cloning procedures. Current Genetics 44,115-123. 

Alfhough we are in transition from the age of S5mfhefic chemisfry fo the age of biofech-nology and gene manipulation, S5mthetic discovery will still play a major role in advancing the therapeutic armamentarium. 

Foster Yes. I think if we are going to understand what melanopsin is doing a comparative approach will be valuable, and this is why Mark Hankins and I have started to work on zebrafish in parallel with mice. Although the transgenics and gene manipulation is not as sophisticated in fish, there are other things that can be done, particularly at the electrophysiological level. As I discussed in the presentation, Mark Hankins and I are keen to pursue this approach. 

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