Considering myself a meso-dinosaur, I would suggest to simply grow your crystals in capillaries and then wick off the liquid and shoot them like this without much handling.
A very efficient and simple trick is to stick a capillary in an Eppendorf tube with the lid on using the Eppendorf tube as a reservoir. I admit this method was used before I was born but works well, not quite high throughput though and need some training in handling etc. As a tip use a syringe to poke a hole into the Eppendorf tube and then wiggle until the capillary fits. Jürgen On Apr 13, 2012, at 1:49 PM, <[email protected]<mailto:[email protected]>> wrote: Hi Prem, In addition to other remarks made: - You could dissolve one or more crystals in water and have mass spec done to verify that your crystals are a complex. It takes many crystals (20-30) to make sure on an SDS-PAGE. You will probably need to silver stain the gel to enhance the sensitivity. And optimize for visualizing DNA (of course protein and DNA would each be control lanes). - Apart from optimizing your DNA length and overhang as suggested, you could also try to see what a detergent does for you. My experience is that they can dramatically improve the crystal quality for protein-DNA complexes. But you first need to know if the crystal consists of both protein and DNA. I am optimistic about the probability. Mark (who apparently is also a dinosaur because he practices room temperature crystallography) -----Original Message----- From: Prem kumar <[email protected]<mailto:[email protected]>> To: CCP4BB <[email protected]<mailto:[email protected]>> Sent: Fri, Apr 13, 2012 5:10 am Subject: [ccp4bb] Crystal behave funny Hi all, I got some Protein + DNA complex crystals (image attached) recently. They are needle shape some times splitted chromosome type crystals. When we pick long needles they bend so much than normal crystal but they dont break. The small needle dissolve very fast as try to open the drop's film. we try to diffract the long needle crystals and they diffract up to 20 A resolution. Any suggestion how to improve those crystal packing. Thanks in advance! -Prem ...................... Jürgen Bosch Johns Hopkins University Bloomberg School of Public Health Department of Biochemistry & Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205 Office: +1-410-614-4742 Lab: +1-410-614-4894 Fax: +1-410-955-2926 http://web.mac.com/bosch_lab/
