CEN sequences on plasmids (yeast)
Posted 22 November 2010 - 09:25 AM
If I transform a plasmid (with an ORI) into a yeast cell the plasmid will replicate once per cell cycle.
How can I be sure that the daughter will get one of this two plasmids?
Therefore I need to integrate a CEN-DNA-Sequence in the plasmid, so the mitotic spindle
can be attached.
-> That is only a consederation of me. I would be happy if you could tell me whether I'm right or wrong!
Posted 22 November 2010 - 10:35 AM
Posted 22 November 2010 - 01:37 PM
selection pressure(auxotrophy) as you just said.
I think the problem would be a mother cell with a lot of these plasmids with my target gene.
This could lead to overexpression and falsify the experiment.
Posted 22 November 2010 - 04:21 PM
Posted 23 November 2010 - 01:48 PM
Then plasmids should be given to the daughter at random.
I think it also depends on the structural condition how much the mother cell is able
to accumulate before sharing plasmids with daughter cells.
I have also read that special Plasmids (called ERC = extrachromosomal rDNA cycles) are
factors that can accelerate the aging process in s.cerevisiae. Besides that, experiments
are supposed to have proven that it is just the number of plasmids which is responsible for
the faster aging. It doesn't matter wether it's rDNA or another plasmid: high amount of plasmid = old cell.
PS: I am not doing any experiments yet. I am just a inquisitive student (and also annoying I think )