TigTigger
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We all use different pipettes....but maybe should try new ones.
Any thoughts about what it could be if not phage??!!! It's obviously immensely frustrating!!
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The overnight inoculations grow fine i.e. inoculating with a single transformant, we then inoculate LB 1/100 with overnight...the cells grow normally for 2+ hours then just as OD595 0.4-0.5 is reached clearing starts and what looks like protein clumps appear in the medium. Earlier today I did a small scale prep (2x500ml flasks), 1 flask lysed before optimal OD reached, other a couple of hours into induction...so something obviously got into that one when I added IPTG!!! The flasks aren't opened at all in the incubator and everything is done using sterile technique.
I inoculated the remaining overnight culture with 1/100 dilution of lysed culture, but no lysis apparent!!! verrrrrry odd!!!!
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Yup and tried using alternative incubators in other locations...it's most puzzling that the cell lysis doesn't occur in every flask...have a suspicion it's something in the lab environment rather than just isolated to an incubator?
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Hello!!
Right we have a leetle problem in our lab and I was wondering if any wise folks out there may have an idea or solution...any thoughts at all most welcome....
Something is killing our E.coli...about 2 hours after inoculation the E.coli start to lyse. Initially this happened infrequently so it wasn't really a cause for concern, but now it happens almost every prep. Sometimes 1 or 2 flasks will survive, so I think can safely rule out problems with reagents. We have tried using new flasks, new stocks of antibiotic, new medium, transformants etc etc. I know that this sounds like a phage infection, serial dilutions of the lysed culture onto plates of confluent E.coli showed no bands of clearing one would expect in the presence of phage??!!!!
Help!!!!!
xx
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NMR DNA substrates
in Biochemistry and Molecular Biology
Posted
Hi
I have a query about ordering DNA substrates for NMR...According to literature for a 1:1 ratio DNA:protein I need to titrate 10mM DNA substrate per NMR reaction. Most of the facilities that offer synthesis of oligos produce uM concentrations. Where can I order these quantites of DNA (in the UK), for NMR (I am also assuming that they need to be HPLC purified?)?
Any help would be muchly appreciated