Difference Between Evolution and Mutation?

[Skye]

Plasmids are pretty simple. Little circle of DNA that can be passed between single celled organisms. The often carry genes that help the organism, this is a problem if it's for say, penicillin resistance. They are very useful for genetics research though, since you can use them to put genes inside bacteria or yeast.

[Radical Edward]

what happens if you stick a plasmid in a eukaryotic cell?

[Skye]

A bacterial plasmid won't be transcribed because the promoter and iniation sites aren't recognised by eukaryotes. So it would eventually get broken down and used for something else. Yeast (a monocellular eukaryote) have naturally occurring plasmids, like the 2-micron plasmid. Plants sort of have naturally occurring plasmids, it occurs in a bacteria, Agrobacteria, which infects the plant cell, and the plasmid is transferred into the plant, integrates into the nucleus. It has a promoter the plant recognises, which leads to the expression of the it's genes. This causes a tumor to form, and the production of opines, which are food for the bacteria.

 

The other alternative is simply to build something like a plasmid with eukaryotic promoters and put it into a plant or animal cell. There are a few ways of getting it into the nucleus, for animals usually mixing the DNA with calcium phosphate, precipitating out the calcium phophate, and inside the crystals are some DNA. These will diffuse through cell membranes and carry the DNA into the nucleus. For plants, where the agrobacteria aren't effective (i.e. for monocots) the most fun idea is coating pellets with the DNA and firing it into the cell.

[Radical Edward]

interesting stuff. Is there no way that the engineered plasmids would work outside the nucleus? not that I can think of any advantage to this, I was just wondering about mtDNA and how that works. Could engineering these plasmids be a possible method of genetic engineering?

[Skye]

Mitochondria are similar in some ways to bacteria, they don't have introns/exons for example, so you would run into some problems.

[Radical Edward]

sorry, I'm not too sure what you are refferring to there. Mitochondria are more related to the archaea than bacteria though I forget the name of the scientist (a woman if I recall) who proposed that modern eukaryotes are colonies of archaea and primitive bacteria.

[Skye]

Lynn Margulis was the woman. As far as I remember, by sequence comparisons mitochondria seem closely related to the small parasitic bacteria. I think the nucleus, and maybe some other organelles, have alot in common with the archaea though.

 

Anyway I meant that the differences between mitochondrial protein synthesis and eukaryotic synthesis would make it difficult to use plasmids carrying eukaryotic genes. Not impossible I guess, but probably not worth the trouble. Mitochondria are membrane bound, and access is about as well controlled as the nucleus. And mitochondria aren't really protein synthesis powerhouses, they only have a couple of dozen genes I think. Probably most illustrative is that I don't think any viruses utilise mitochondria.

[lucaspa]

The other alternative is simply to build something like a plasmid with eukaryotic promoters and put it into a plant or animal cell. There are a few ways of getting it into the nucleus, for animals usually mixing the DNA with calcium phosphate, precipitating out the calcium phophate, and inside the crystals are some DNA. These will diffuse through cell membranes and carry the DNA into the nucleus. For plants, where the agrobacteria aren't effective (i.e. for monocots) the most fun idea is coating pellets with the DNA and firing it into the cell.

The most common method is electroporation.