Is there anyone out there who works in a high-throughput crystallization lab that would be interested in running some experiments to see how well "random" microseeding / rMMS works when it is used routinely (i.e. as soon as the first crystals stop growing)? I have in mind a very simple protocol and a simple way to measure its effectiveness for getting more structures.
The reason I am asking is that the obvious thing for both William and Anamika to try is rMMS, as has been suggested many times on this bb. I don't understand why everyone doesn't know about and use this method, which was published seven years ago. I think part of the reason is that - although lots of small labs use it - as far as I know none of the high-throughput centers use it routinely at an early stage. I've approached several H.T. groups about this but none of them were interested in giving it a try. If anyone from a high-thoughput lab is interested I'd be very happy to stop by and give a seminar with all the theory and practical details, plus put forward some simple ideas for testing the effectiveness of the method. Please let me know off-board. William and Anamika, I would make a seed stock with the crystals that you have and add it to a couple of random screens. References and practical info are below. (B.t.w. it very often works well with sea urchins, William.) Best wishes to all, Patrick __________________________ Refs etc: 1. Galina Obmolova reported in 2011 that 38 of her 70 structures were determined using MMS. That included 70% of the complexes. See slide 5 of http://hamptonresearch.com/documents/ramc/RAMC2011_T11_Obmolova.pdf 2. The original publication by D'Arcy *et al.* (based on Stoddard's work): http://scripts.iucr.org/cgi-bin/paper?S0907444907007652 3. Our (slightly "heavy") article in Crystal Growth and Design about how to adapt the method to different situations: http://pubs.acs.org/doi/abs/10.1021/cg2001442 4. A fun project that we were involved in where "random" cross-seeding worked brilliantly: http://scripts.iucr.org/cgi-bin/paper_yard?wd5214 5. Our latest recommendations for the method: http://www.douglas.co.uk/MMS_proc.htm On 11 April 2014 01:50, william lee <[email protected]> wrote: > Dear All, > > > > I am currently working on a ligand bound protein complex. From my initial > crystallization screens I have identified a condition which gives me sea > urchin like crystals. I managed to repeat these crystals in my optimization > conditions, in fact I can see crystals in all the wells but there is no > significant difference between them. All conditions give me similar size > and amount of crystals. To confirm these are protein crystals or not, I > tried expose the crystals to X-ray beam (in-house). Good news is that I > have no obvious salt diffraction at high resolution, but the bad news is my > low resolution diffractions are not really believable. In addition, these > crystals seem to be quite easy to separate into needles but they are too > small to tell if they do crack like most the protein crystals. > > > > I have attached a picture of my crystals and the diffraction pattern I got > at low resolution. > > > > I am hoping if anyone can suggest me on how to improve or change the shape > of these crystals if they are genuine protein crystals. > > > > > > Many thanks > > > > Kind regards > > William > -- [email protected] Douglas Instruments Ltd. Douglas House, East Garston, Hungerford, Berkshire, RG17 7HD, UK Directors: Peter Baldock, Patrick Shaw Stewart http://www.douglas.co.uk Tel: 44 (0) 148-864-9090 US toll-free 1-877-225-2034 Regd. England 2177994, VAT Reg. GB 480 7371 36
