Purchase or make competent cells? - (Apr/18/2007 )

I'm curious as to whether most of you are purchasing or making competent cells, and what the trade-offs are. It's strange because I see a lot of discussion about how to make competent cells in these forums, which makes me think lots of people are making their own, but also see a lot of companies offering a large variety of competent cells, which makes me think there's a lot of demand for purchasing them.

Thoughts?

I believe most labs make their own competent cells, it is simply to expensive not to do so.

The trade off is that home made competent cells which are very cheap are far less efficient then company cells.
(Home made) 10^6 compared to (company) 10^10

For most situations home made cells are good enough, but there are those hard ligations, like ligating very large plasmids, BAC, PAC and very DNA fragment together. The higher efficiency of company cells is need in such a situation.

The reason for this inefficiency is that we don't invest substantial time and effort to improve the generic hand me down protocol that we use. (which might be surpassed by better methods decades ago). And if some brave soul has done the optimisation work, we as a community have been very disorganised and been unable to enshrine said work and transmit it efficiently to each new generation of PhD students.

Companies have capitalised on this malaise and/or disorganisation to basically charge blood for competent cells which differ only by the protocol used in their processing.

Getting competent cells from companies prove one thing which is they do provide quality control over their products and thus their transformation efficiency is extremely high. But the down part is it is very very expensive to buy those competent cells.

On the other hand, making own competent cells are pretty tedious. But it is very cheap to make it. Of course, the quality control isnt that good. Sometimes I got extremely low transformation efficienty (certain tube). But I guess as long as you are able to transform desired plasmid, I don't see any problem with that.

I have spent the time to optimize competent cell preparation, and to compare the bewildering array of possible techniques. We routinely now make cells as good as commercial chemically competent cells, as measured by standard transformation tests. We make them in high volume, about once every two months. See here:
http://openwetware.org/wiki/TOP10_chemically_competent_cells

I have not invested my time to do it by myself, but I did squeez it to the maximal usage. I took some advice from Bioforum and cut the amount of bacteria used for each tranformation. Instead of 50 ul as recommended, I use only 10 ul. For my purpose, I got more than enough colonies from each transformation, which makes the purcahse less painful. hehe. Poor man's trick.

Thanks for the protocol I will be sure to give it a go. Just curious though - why grow the bacteria on SOB plates first? Are LB plates ok for a substitute or should I stick to SOB?

we make our own competent cell, as other said it is tedious. now i have only 4 tube of competent cell, i need to make it soon, i already made the media but when i think to make the cell i really feel tired......

It's probably not important to use SOB plates for the seed stocks, but since this is essentially a once in 10 years event, it is hardly much of a burden.