Briefcase Sized DNA Analysis System

An anonymous reader writes "Japan's NEC Corporation along with Aida Engineering have developed a briefcase-sized DNA analysis system that enables the police to perform comprehensive DNA testing at crime scenes in as little as 25 minutes. The same test would take at least a day to a week (if re-testing or conformation is required) in the lab. The system is compact enough to be carried to crime scenes or other locations where quick DNA analysis is required, making it the world's first portable DNA analysis system."

Comments

processing time claim is very optimistic.

[mauthbaux]

From TFA:

The compact unit can be used to:
(1) take cell samples,
(2) extract the DNA,
(3) perform polymerase chain reaction (PCR) amplification to generate copies of the DNA,
(4) perform electrophoresis to measure the spacing between DNA bands (to create the genetic fingerprint), and
(5) perform short tandem repeat (STR) analysis to create a unique genetic profile for the individual,"

As I'm currently a grad student in biotechnology (and am performing similar processes in the lab), I feel compelled to respond to their claims on processing time. Taken step by step;
1: simple enough, although some cells are more suitable to DNA work than others.
2: the main obstacles in extracting DNA are proteins and prokaryotic contamination in the sample. DNA is almost always complexed with proteins like polymerases and histones. These proteins effectively prevent the DNA from migrating through agarose or acrylamide; the resulting electrophoresis bands would be almost meaningless. Prokaryotes are pretty much ubiquitous. The problem is that they carry their own DNA which can confuse results, and they carry endonucleases which chop apart most any DNA they come in contact with; destroying the reliability of the gel electrophoresis. Endonuclease digestion of DNA is standard fare for genetics, and I'm assuming that it's performed here, but the contamination of unknown endonucleases from uncharacterized bacteria causes problems regardless. Time required to separate the DNA from the proteins: 1 hour at best.
3: PCR incubation time depends on the length of the DNA chains being amplified, and the initial size of your sample. 10 minutes would be a best case scenario, and that's with ideally sized DNA fragments (whole-genome DNA is far too large), and a large initial sample (not likely).
4: As mentioned previously, protein contamination can make the electrophoresis results almost unreadable. Furthermore, moving that much DNA through a gel in such a short time requires very high voltages. The banding which results from high voltages is generally very blurred, making the 'fingerprint" unreadable. Moreover, whole-genome DNA doesn't really separate into bands; it makes big long smears, so standard staining practice is useless for diagnostics. The last gel I ran with genomic DNA (corn in this case) required about 45 minutes, and that was a small gel using high voltage.
5: STR analysis is touchy. Basically, you use a radioactive or chemoluminescent probe on both the genomic DNA, and a DNA with known STR lengths and compare how bright the sample is compared to the standard. An accurate reading requires a fairly precise estimate of the amount of DNA in your sample; a measure that usually requires a well-calibrated photospectrometer that also needs time to warm up and be calibrated. To further complicate matters, your DNA is in a gel. Getting the tagged probe into the gel (or getting the sample DNA back out of the gel) so that annealing can occur takes time. A southern blot (process involving the removal of DNA from a gel) is usually allowed to run overnight. After annealing takes place, the extra probe molecules are washed away. If excess stray probe is allowed to sit around, or if the annealing isn't complete in the first place, the measurement becomes unreliable. The minimum time I would think feasible for this step would be an hour. In a lab, the labeling alone is normally a 3 hour process. Accuracy would suffer tremendously as time decreases.

So yeah, in conclusion, their time frame for getting results is obscenely short. Severly truncated time frames produce equally severe errors. I don't personally know any scientist who would vouch for the validity of these results.

Re:processing time claim is very optimistic.

[flushingmemos]

I imagine they can do it in the time they say. I also imagine the results are very simple, like looking at one STR sequence and counting how many lengths of it are in the person's genome in a process similar to qPCR, less RFLP/southern, as parent seems to think. Despite what TFA might imply, I don't think there's endonuclease digestion involved. I may be wrong, and they could have a really, really fast breifcase thermocycler making this work. Maybe, doubt it.

I'm not any kind of STR expert, but from cribbing Wikipedia http://en.wikipedia.org/wiki/Short_tandem_repeat, here's my impression of what's going on with this kit:

1. get cells = blood and semen. yum. In fact, I'd infer this kit is probably a "semen-only" deal in practice, which makes isolating the DNA that much easier, since semen is largely DNA.
2. isolate DNA. Do it yourself, kids! (http://learn.genetics.utah.edu/units/activities/extraction/) 2-5 minutes with a kit.
3. PCR. Here's where things get interesting. What are their primers? I think they're using 5-10 nucleotide STR sequences that are already conjugated to a fluorescent dye. Since STRs for human identification use are just, according to wikipedia, 4-5 STRs (10-50 nucleotides) long, each cycle can probably be as short as 30 seconds. With ramping the temperatures we can call that 1 minute per cycle. How many cycles do we need? 10 cycles gives us 2^10 copies of the original STR, that's (biologist math)1000 copies(/biologist math). Add 2 minutes for our hot-start polymerase, and that's 12 minutes for PCR. Whoo! It well may be less, i.e. shorter elongation, fewer cycles. This is where they're claiming to save time, so who knows.
4. electrophoresis. Undoubtedly capillary, you can see it in the photo (at least I can), and since we're looking at stuff that's only 75 nucleotides max, can be done very quickly. I don't really know capillary gel electrophoresis, but it apparently kicks the shit out of slab gel electrophoresis: http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=330505. We'll call that 10 minutes, lol. Could be a bit more? Balances with the PCR.
5. Of course, we added a 10-100 nucleotide standard conjugated to a different fluorescent species from the primers (i.e. the primers glow green and the standard glows red), so we can use our shitty little built-in 2-wavelength spectrophotometer to see where our unknown sample's bands are.

And now we have our data! And that only took... 25 minutes! Of course, this isn't a full-blown RFLP, like parent seems to assume. But just for doing a quick-and-dirty count of STRs, this could work. That's how I'd do it. Maybe I just invented a competing type of kit, lol. In any event, looking at the picture, I get the feeling their pipettes are crap.

Note this doesn't show how many repeats of a given legth the accused has, so the asshole could have 3 5-repeat ones and 2 4-repeat, and the machine would show that as being the same as a person with 1 of each. Also, they may use more fluorescent dyes to look at more STR sequences without too much difficulty. But in general, the samples will be unclean at best, total crap more often than they'll like to admit, and, in the end, only good as a blood-type-and-then-some test. How juries will react to this, I don't know.

To get even farther from parent, the real threat to your privacy is coming from gene chips, the next generation of sequencing technology. This kit is comparatively rudamentary, and obviously expensive. Yet more overhyped crap, whee!