davison@bnl.UUCP (daniel burton davison) (12/14/84)
[this line does not contain a leading white space] > alexis@reed says: > "...and the restriction in the number of copies in *higher* > organisms is no doubt artificial." (emphasis added) There have been a number of side points in this discussion, which started as consideration of a self-reproducting organism. Some of these contain very subtle errors in logic but I will not pursue them here. [anyone who is interested can contact me by mail at the address below]. I initially challenged the assumption stated above that the restriction of the number of copies of a gene in higher organisms is artificial, i.e. a result of synapsis-dependent mitosis/meiosis. >alexis@reed responded by shifting the argument to lower organisms: > Bacteria generally have more than one copy of their plasmids. > Mitochondira...also have...a variable number of copies of their > genetic material. Some bacteria shuffle their genetic material > by...[passing]...a RANDOM LENGTH [of chromosomal DNA]. Bacteria do not generally have more than one copy of their plasmids unless strong selection is present. The F plasmid, R factors, various cryptic plasmids are all present in usually one copy. If an R-factor encodes a useful antibiotic resistance gene, the number of copies will go up, **but** many more copies of the resistance gene will appear on the R factor by unequal recombination and disappear when the selection is removed. Incidentially gene duplication is the basis of low-level ampicillin resistance in E. coli (from the chromosomal beta-lactamase). This effect also works on genes such as lac. [I'll provide references to anyone who is interested]. The random length of DNA passed by F or R factor mediated conjugation is either incorporated by recombination or destroyed. Try a three factor cross some time. The bacterial plasmids which do have multiple copies (the colicin plasmids) are in general non-deleterious to the host and very non- essential. As an example of a high copy number plasmid the pBR322 cloning vehicle (colE1 and some other odds and ends) will live in about 10-20 copies per chromosome. But if you put some genes on it or remove selection all bets are off. Consider the problems of judith Zyskind et al. on trying to clone the origins of replication of various Enterobacteriaceae. Also, in an exponentially growing culture of E. coli, each cell has four chromosomes in various states of completion-hence the copies/ chromosome mentioned above. You've really got 4 bacteria there. Mitochondira and chloroplasts have one copy of their genome per organells --ask anyone who isolates mt or cp dna often. > In higher organisms...But I am pretty sure that such restrictions result >from the fine tuning of other mechanisms that have come to depend on a > constant amount of DNA. This last phrase troubles me. I can't think of any mechanism of gene or developmental regulation known or suspected which requires a constant "amount of DNA". Lillies have several (100x ?) times the amount of DNA per cell as a human cell has. Not even the length of the cell cycle is proportional to the amount of DNA; new origins of replication are added. The key here is that we are mixing apples and oranges, or if you prefer genes and nuclei. Yes, certain genes can exist in hundreds of thousands of copies outside of the chromosome (dihydrofolate reductase is an excellent example); these are gradually lost with the expected kinetics when selection is removed. So, are there any mechanisms in procaryotes which limit the number of copies of the *genome*? The answer from Enterobacteria is yes: try maintaining two enterobacterial origins of reolication in one cell, even with strong selection. At the sub-genome level, such a mechanism exists; and every plasmid family that I've ever heard of has incompatibility and copy number control genes. In fact the incompatibility mechanisms are quite ingenious. I had 1.5 years of work blown out by incompatibility between the plasmid I was making and a cryptic plasmid in Salmonella typhimurium LT2; even ****strong*** selection didn't maintain it; the cells just died. Slowly. I'll be glad to provide references to whoever wants them. dan davison PhD, 1985, molecular genetics arpa: davison@bnl uucp: ...decvax!philabs!sbcs!bnl!davison