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You’ve cultured your cells and completed your treatments, now it’s time to harvest them and proceed to the downstream effects. Cell lysis is the crucial stage that determines if your experiment has a chance of producing the data that you have been waiting for. Part of the starting biological material is inevitably lost on each…
Put in the time now to optimize protein solubility so you don’t have to suffer later! With this in mind, here are some considerations and tips for selecting the optimal conditions for recombinant protein expression and purification.
Ever wonder what those small colonies, like satellites, surrounding a larger E. coli colony on your LB with ampicillin plates were? Or why, when you picked that colony, it never had the plasmid you just transformed? Well, it’s because those satellite colonies are “protected” from the ampicillin by the big colony. Read on for more… Ampicillin…
In the sci-fi novel Terminal World by Alistair Reynolds, a planet consists of zones with defined characteristics of matter interactions on a subatomic level. These conditions permit different levels of technology sophistication in various zones. For example, in the “Steamville zone” nothing more complicated than steam engines works – electronic schemes fuse irreversibly. Something like…
As a protein biochemist and a Ph.D. student, I was given the task to express a eukaryotic protein in a bacterial system. And to say that I was having a hard time would be an understatement. It took me many PCRs, cloning and transformations to get to the right construct that would eventually express the desired…
Dear Aunt Yersinia, A very annoying postdoc in our group keeps telling me off for spinning E.coli at 13K in a tabletop centrifuge. The postdoc claims that high speed damages cytoskeleton and this will reduce my transformation frequency. But I don’t believe her as the cells are cushioned by water during centrifugation. Can you tell…
Bacterial cultures may be much easier to grow than mammalian cells, but if your yields are suboptimal there are plenty of parameters to play with. Here we list a few of the things you should consider to maximize your culture growth. Shaking speed Shaking is performed to allow aeration of your culture, which is of…
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