New DNA Fragment Sizing Technology - help understanding the scientific value, if any (Nov/03/2005 )
Please forgive the random email. I am trying to reach scientists that may be able to help me evaluate the scientific value of some technology Los Alamos National Laboratory has developed for DNA fragment sizing. The technology offers much better resolution than gels for large DNA fragments - the dynamic range is from 5000 up to 500k bases - and requires very small amounts of sample (on the order of microliters or only several thousand fragments in the sample). The Los Alamos technology could be turned into an inexpensive instrument that would analyze a sample in a few seconds and would cost only a few cents per sample to operate.
I am not familiar enough with this area to know if there any subset of people doing DNA fragment sizing that might be excited by any capability (small sample size, rapid sample analysis, or very large fragment sizing). I would be eternally grateful for some help. Or if you know of someone that might be able to help, that would be useful as well.
Hi, I am not sure about a specific use for this technology, but it sounds like the type of thing that people will be looking for to aid in genome wide analysis of things...
potentially being able to see any size DNA on the order of thousands of fragments and rapid analysis may be useful, but (and this assumption may be wrong) I would suspect that your sensitivity arises mostly from the large size of the DNA, so that the number of fragments required for visualization would increase proportionally to the size of the DNA?? Anyway, if this is not true then you could sell this technology based on the increased sensitivity of detection which is something we are always looking for...
I hope that my rambling is useful, I just cant think of a specific application at this point...
good luck
also interesting from my point of view would be, how easy visualisation would be? at the moment most staining proceedures involve harmful substances, so if your technique does not, it's a definite plus. what about recovery of a fragment of defined size? that is also an important feature
That could also be interesting for quite a number of appication of whole genome genotyping, to replace the long and hard step of PFGE. But it could be even more interesting if slightly smaller fragments could be detected (say starting in the 1000bp range).
I would definitely use this kind of thing!
I would definitely use this kind of thing!
I agree with that! Would be very useful for a laboratory like where I am, that does a lot of bacterial typing.....
Cheers,
Stegger
Kersten,
Sorry for the delay in following up on this thread. The technology works by single-molecule detection. So we bind a ubiquitous dye like Sytox Orange to the DNA fragments, stretch them out in a thin flow stream so that only one labeled DNA is in the field of view at a time, and then excite the dye and watch the intensity of the emission. Emission intensity tells us how many Sytox Orange fluorophores which correlates to the size of the DNA since larger molecules have more dye stuck to them.
The amount of stain used is trivial - one order of Molecular Probes Sytox Orange has lasted for years.
You can sort out fragments of a particular size however they will have the dye stuck to them so they have to be useful in that state. Different dyes are possible, of course.
There is also the potential for finding fragments with specific sequences in them. If a second dye is bound to a probe that hybridizes with a specific sequence, then the system can be configured to look for emission at two wavelengths. Emission from the ubiquitous dye means there is DNA in the FOV and simultaneous emission from the probe fluorophore means that the DNA fragment contains the sequence of interest. If that has any utility.
I'd love to talk more directly; please feel free to email me at elling@lanl.gov
Why do you need to streach out the DNA if you are only measuring the emission intensity? Also do you have a paper or website about this technology?
Thanks
Rob