Why does NaCl aid the growth of e-coli?
I know that it does! But noone can tell me WHY! Also, this is technically two questions- is e-coli capable of taking up a linear plasmid? As in one that has been restricted by an endonuclease and not reannealed. Please help!
Answers:
NaCl needs to be at the correct concentration, as it is needed to ensure that enzymes etc. work correctly. It allows the cells to have the correct concentrations of sugars etc by influencing water entry by osmosis.
A cell can take up a linear plasmid, but probably won't be able to propagate more copies if is isn't annealed, unless the correct DNA Ligase enzymes happen to be available in the cell.
? Under what conditions? Maybe you are just adjusting the osmotic pressure of the medium. Above 0.6 M (if I remember correctly) E.coli cells don't grow.
Yes they can take up a linear plasmid and if the ends are phosphorylated they will ligate it and proliferate happily ever after.
Usually after restriction I dephosphorylate my vector (especially if using a single restriction enzyme) to minimize background from re-ligated plasmid.
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Answers:
NaCl needs to be at the correct concentration, as it is needed to ensure that enzymes etc. work correctly. It allows the cells to have the correct concentrations of sugars etc by influencing water entry by osmosis.
A cell can take up a linear plasmid, but probably won't be able to propagate more copies if is isn't annealed, unless the correct DNA Ligase enzymes happen to be available in the cell.
? Under what conditions? Maybe you are just adjusting the osmotic pressure of the medium. Above 0.6 M (if I remember correctly) E.coli cells don't grow.
Yes they can take up a linear plasmid and if the ends are phosphorylated they will ligate it and proliferate happily ever after.
Usually after restriction I dephosphorylate my vector (especially if using a single restriction enzyme) to minimize background from re-ligated plasmid.
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