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    How Did Science Determine The Body Was Osama Bin Laden? PCR
    By Hank Campbell | May 2nd 2011 01:57 PM | 27 comments | Print | E-mail | Track Comments
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    With Osama Bin Laden dead, conspiracy theorists will find a way to say it isn't him at all.   Sure, a 6'4" thin guy can be replaced by decoys rather easily but science has come a long way since the September 11th, 2001 attacks that took Bin Laden from being a famous terrorist to being infamous - though given recent developments in the middle East and Africa, Bin Laden has ironically done more to promote democracy in the region than anyone, since the establishment of two democracies in retaliation for regional support for Bin Laden has had a domino effect.

    Visual identification by a SEAL team (not seeing his face but actual facial recognition technology), and confirmation by a local is good, but not always enough to satisfy everyone.  

    Here's how they were able to determine the body was really Osama Bin Laden: PCR.  

    Polymerase chain reaction (PCR) means scientists copy a stretch of DNA (deoxyribonucleic acid) using heating and cooling and then add fluorescent dyes to compare specific spots on that chunk of DNA with a sample from a relative or an older sample.  PCR has been in existence for decades, since 1983, and Kary Mullins received a Nobel Prize for it in 1993, so it has stood the test of time.

    A gene's building-block sequence is the order of appearance of 4 deoxyribonucleotides within that stretch of DNA - Adenine, Thymidine, Cytosine and Guanine, which are the A, T, C and G you often see. The arrangement of the letters is the gene sequence. DNA is double-stranded (except in some viruses) and two strands pair with one another and each letter in a strand will pair with only one kind of letter across from it in the opposing strand: A  with T and C with with G, for example.

    The first step for PCR is to create "primers" about 20 letters-long, using each of the 4 letters, and a machine which can link the letters together in the order desired by adding one letter-at-a-time to the machine (DNA synthesizer).

    PCR

    For most labs PCR takes about two weeks, though it can be done in a day.   Contamination is often the biggest concern but since they brought Bin Laden's body back with them, they didn't have field conditions to worry about.

    Want to do your own PCR?   SUNY Plattsburgh in upstate New York has a nifty tutorial.

    After you heat the DNA to separate the paired strands and your primers, as mentioned above:

    1. Take three tubes and mark them 10-2, 10-4 and 10-6. Put 199 ul of H2O in the 10-2 tube and 990 ul the 10-4 and 10-6 tubes. Put 2 ul of your original plasmid in the 10-2 tube and mix well. Put 10 ul of the solution from the 10-2 tube in the 10-4 tube and mix well. Put 10 ul of the solution from the 10-4 tube in the 10-6 tube and mix well. Each of these is a 100 fold dilution.

    2. Label 2 PCR tubes (the 0.65 ml tubes provided) 10-4 and 10-6.

    3. Set up the following reactions in each of the tubes (I will be adding the buffer mix and Taq polymerase):

     34.5 ul H2O 1.0 ul diluted DNA solution 14.0 ul Buffer mix (buffer, primers, MgCl2, and dNTPs) 0.5 ul Taq polymerase 50.0 ul Total

    4. Place these three tubes in the thermocycler.  You have one of those, right?After you have run the agarose gel, note the extent to which you diluted the DNA in the reactions. Assuming that your original plasmid stock was approximately 1 ug / ul (which is probably a good estimate for most of you), the 10-6 dilution had a concentration of 1 femtogram / ul. This means that your 10-6 PCR reaction only has about 1 picogram of DNA template. (Imagine - one picogram!) For this reason, on the agarose gel you should only see the amplified fragment and not the DNA from the original template.

    Comments

    '...but science has come a long way since the September 11th, >>2011<< attacks that took...'

    You might want to get that fixed.

    Hank
    Gah!  Thanks.  The advantage of having no editors is...well, we look dumb before someone tells us.
    "Contamination is often the biggest concern but since they brought Bin Laden's body back with them, they didn't have field conditions to worry about."

    Just curious about this sentence... they brought Bin Laden's body back with them to... where, exactly? A nearby laboratory? A lab at a military base? The Navy ship? That's as far as the body got.

    Hank
    Given what they said about the planning for this - and they seem to have planned it extensively - a lab was easy to set up in advance.  They have to have known about doubts otherwise so verifying the identity was key.

    Wetwork in a kitchen is tough but not with unlimited government money.
    While a thermocycler is cheap enough and portable to carry around, setting up an ABI capillary electrophoresis analyser on a Navy ship would be expensive and rather odd for a one off test.

    Since I estimate a minimum four hour turn around time once a sample arrived on the lab bench and DNA results were being announced almost immediately after Obama's announcement - I think it is safe to conclude that the government is lying about DNA confirmation.

    Hank
    It is more likely, conspiracy theories aside, that he was instead killed earlier than announced.
    //It is more likely, conspiracy theories aside, that he was instead killed earlier than announced.//

    This is the only way I can figure out that they got good quality results prior to the announcement. Of course, they could very easily have had a test in process and announced that it was confirmed instead of being confirmed. We've seen how sloppy the media can be when they're in a hurry. I hope they make the test protocol and results public, thereby eliminating the need for gruesome photographs. (Oh, wait, the public still wants to see blood :P)

    Hank
    The public does indeed and a lot of it has to do with party affiliation and media - the lack of peace protest coverage since the 2008 elections tells anyone in the middle something is wrong with how things are portrayed now versus then.   If Bush were president not only would 30% expect to see a body but there would be calls for a war crimes trial because OBL was unarmed.   Since Obama is a Democrat, a different 30% think he fabricated it to look tough enough to be president and distract people from the economy and they are making fun of him for not wanting to arrest OBL instead.

    I suppose you can fake a DNA test too - but it is a lot harder.
    No need to bother faking it when all you need to do is have John Brennan say we have done a DNA test and 300 million Americans obediently chant after him "we have done a DNA test"

    The Stand-Up Physicist
    I built my own PCR machine in 1988, the Dr. Gumby Go-Round. There were 3 heat blocks sitting on a turntable, with an arm that went up and down that held the eppendorf tubes (0.5 ml, the small ones). The arm could also hold on to a green Gumby, but he had a tendency to knock over a Pokey. It used an Commodore 64 as the controller. The machine was used to study RNA recombination in the polio virus after I built the machine and left the program. Not a battle hardened machine, so would not have worked for this job.


    The trickiest part: getting electricity to the middle of the turntable. The 4 plugs looked standard, but I had to use brushes underneath to avoid twisted wire. I wonder if that was really unsafe, given that the box was made of metal...

    Doug
    Hank
    That's not the quirkiest thing a biologist here has done in the lab:


    That cool shirt he is wearing is a one-off.  He is literally the only person in the world with it, because we order one of each when we do new shirts, to see what the quality is like, etc. and Mike White got that white golf beta version.  It never went into production, we did long sleeve buttondowns instead.
    The Stand-Up Physicist
    OK, he wins a quirky, but did he do any biochemistry with Mr. Baseball? I know my PCR machine was running a full 5 years after I had been set out to pasture. Dr. GGR got a Ph.D. for a hard working grad student. Perkin Elmer was charging $10k with a 6 month delay for a PCR machine. I got to go into machine shop rehab for a month (the wise old machinist would start the day, "Well, you could do it that way, but this would be better").
    "It is more likely, conspiracy theories aside, that he was instead killed earlier than announced."

    Well either way the government is lying. Why should they not announce when he was killed?
    Personally I don't doubt that they have a corpse that was Osama bin Laden [however that he may have passed onto the next world] - frankly I don't care one way or another, but I think it is more likely to be true than not.

    I am just certain that they have not done a DNA genotyping - either in Bagram or on board an aircraft carrier. That is my only point. A wikipedia summary of the various accounts is as follows: "ABC News, the Associated Press and The New York Times have reported that bin Laden's body was identified by DNA testing,[10][100][101] using tissue and blood samples taken from his sister who died of brain cancer.[102] Reuters reported, however, that DNA test results will be available in the next few days and that bin Laden's body was identified using facial recognition techniques.[103] One of bin Laden's wives identified the body.[104] ABC News stated, "Two samples were taken from bin Laden: one of these DNA samples was analyzed, and information was sent electronically back to Washington, D.C., from Bagram. Someone else from Afghanistan is physically bringing back a sample."[44]"

    At very least reports appeared in the media of a positive DNA ID before they had time to run the test. Again, the issue here is not disputing if OBL was killed or not, just amazed yet again at the fact that Americans seem so complacent and accepting when their government tells them bare-faced lies.

    Since the questions are so simple - where is the sequencer, why is it there, was it just set up for this operation - why does the media not ask them?

    I don't know exactly when the first mention/reports of DNA testing came out relative to the death, but it was something like 6-7 hours between the time of the raid and the president's speech, and I don't think the DNA testing was mentioned in the speech, but sometime after. That would be a really quick turnaround time, but not impossible. I wouldn't be at all surprised if they have a DNA lab/labs somewhere in Afghanistan already, not to mention that, like Hank said, they seemed to have done extensive planning over the course of several months for this raid, so setting up a lab just for this purpose would be simple enough.

    It's also possible the first test wasn't a traditional run. There's lots of research that's been done on faster analysis procedures. It really wouldn't surprise me that much if they used something like this for a quick "screening" test, and then sent the samples on for analysis with a more thoroughly validated system (which would match the news stories of DNA already having been confirmed, and still waiting for results a the same time). I've read papers of full 13-locus multiplex PCR reactions done in under 20 minutes, and post-PCR cleanup leading to time-of-flight mass spec can get a full profile in around the same time. If they did mtDNA (which sounds likely given the mention of comparison to samples of Osama's sister) I'm sure similar types of techologies/procedures are out there, though I'm not familiar with them.

    Gerhard Adam
    ...just amazed yet again at the fact that Americans seem so complacent and accepting when their government tells them bare-faced lies.
    You're quite quick in asserting that these are lies, despite the obvious evidence that you don't know what you're talking about regarding the DNA analysis itself.

    http://homelandsecuritynewswire.com/fast-dna-analysis-law-enforcement-unveiled
    http://pubs.acs.org/doi/full/10.1021/ac101891k
    http://www.ksat.com/news/27761269/detail.html

    While I'm certainly not naive enough to believe that governments don't lie, I'm even less impressed with the "experts" that come crawling out of the woodwork when it comes time to advance their respective conspiracy theories about how governments are always wrong.  I simply wish that all that "intelligence" would be used when it comes time to actually elect these people.
    Mundus vult decipi
    So did the Navy Seals use a Zygem microfluidics chip based DNA test? No, because the system is still in prototype and hasn't been released. Let me just say I have scored more STR genotypes than you have had hot dinners.

    Unless there is a genotyping facility at Bagram airfield - and the latest I could find was in 2008 that they were still sending samples back to the United States - then no DNA testing has been done.

    And lets face it, why would they bother? Americans are so stupid and cowed they won't even ask their government to tell them where the genotyping took place.

    Gerhard Adam
    Americans are so stupid and cowed they won't even ask their government to tell them where the genotyping took place.
    Since you obviously like to live in your own little fantasy world, perhaps in that version of the universe the facts align in the manner you suggest.
    Let me just say I have scored more STR genotypes than you have had hot dinners.
    Like most of what you've posted, I don't believe this either.
    Mundus vult decipi
    So did the Navy Seals use a Zygem microfluidics chip based DNA test? No, because the system is still in prototype and hasn't been released. Let me just say I have scored more STR genotypes than you have had hot dinners.

    Unless there is a genotyping facility at Bagram airfield - and the latest I could find was in 2008 that they were still sending samples back to the United States - then no DNA testing has been done.

    And lets face it, why would they bother? Americans are so stupid and cowed they won't even ask their government to tell them where the genotyping took place.

    Hank
    So did the Navy Seals use a Zygem microfluidics chip based DNA test? No, because the system is still in prototype and hasn't been released.
    I am hopeful some of our taxpayer money goes toward having special ops with some technology before it is released to 16 million academics in schools nationwide.
    Yeah but your hopes are not the issue here.
    Either the Navy Seals had special prototypes or they didn't. We both know they didn't
    Either there is a genotyping facility at Bagram or there isn't. As far as I can tell there isn't, or at least there wasn't as of 2008.

    So if the genotyping wasn't done at Bagram you are looking at least at another 24 hours to transfer a sample state side and run a test. Again the question becomes was this done or wasn't it.

    And again, we both know the answer to that.

    Hank
    Yeah but your hopes are not the issue here.
    Either the Navy Seals had special prototypes or they didn't. We both know they didn't
    We don't know anything.   You make unsupported speculation based on the idea that everyone (except you) is a liar.  It's conspiracy kook nonsense unless you have some evidence.    I accept there are a fringe group of Republicans who will believe anything because Obama is a Democrat just like  idiotic Democrats invented nonsense about Bush and stuck to it, but it doesn't belong on a science site.  It's evidence or nonsense and we both know which side you fall on.
    Even if they didn't have advanced/protoype equipment, they would have had enough time using existing, validated, on-the-market kits/procedures and equipment if they're well prepared,ready to go, and have dedicated personnel and instruments. Figure about two hours for extraction using something like DNA IQ which allows you to skip quantification, about three hours for PCR, then another hour for CE. (Alternatively something like ID Direct is an option which skips extraction completely, which probably cuts about an hour off iof this). That's a total of six hours, and my figures are probably a bit liberal (I've never used DNA IQ personally, but it looks like it could be done in a good deal less than two hours; PCR w/ Identifiler Plus should really only take about two and a half hours, and a CE run typically only takes about 40 minutes on an AB 3130). So we're looking at six hours for the lab work. Once that's done, if you know the profile of the person you're looking for (which would have been done in advance in this case) and you have good clean single source samples (which you should in this case, given they were taken directly from the body), data analysis is a snap.You might spend a few minutes checking out your ladders, correct sizing, viewing your controls, etc. but after that's done it's pretty much just a matter of seeing if the numbers all match, which any first grader could do in about five minutes.

    Of course, that's all assuming they used nuclear DNA analysis. I've seen some mention of comparing with his sister's DNA, which would indicate mtDNA instead. I'm not too familiar with mtDNA analysis, but I understand it is a lengthier process, so if that's the case, I don't know. If they didn't use mtDNA, there's also a question of what they were comparing the profile generated from the body against. It's possible they have a sample of some sort from hism obtained previously, but given that he hasn't exactly been easy to get ahold of, that's probably not the case. If they could get samples from enough of his children and, especially if they also got samples from their mothers, they could have reconstructed a (partial, at least) profile with those. I'd be most interested to hear about this aspect, I think, but I'm sure we'll never hear those sorts of details.

    As for whether they have a lab at Bagram specifically, I don't know, but here's a quote from an article on the US Army website (http://www.army.mil/-news/2010/05/27/39956-shining-light-on-battlefield-...).
    "Until recently, investigators also had to ship all of their evidence back to the States for processing at USACIL. But in 2005, the first joint expeditionary forensics facilities were established in Iraq to process fingerprints, firearms and even DNA collected downrange. The JEFFs have since been deployed to Afghanistan as well, with additional labs ready to deploy on an as-needed basis."

    So not only do they have forensics labs in Afghanistan, they have additional labs ready to deploy on an "as-needed basis." Again, given the amount of planning that apparently went into this mission (including, from at least one source I've seen, a reconstruction of the OBL compound for training), it's not at all out of the question that they set up a special purpose-built lab somewhere nearby for this mission.

    //At very least reports appeared in the media of a positive DNA ID before they had time to run the test.//

    Is it possible that samples had been submitted for confirmation, and the media took that to mean that confirmation had taken place? You know how journalists are with science. (I love journalists, I really do. But we didn't like sitting through their Brit Lit classes, and they yawned their way through Bio for Poets.)

    Larry Arnold
    We don't know, we never will know because there is no independent verification outside of the people in whose interest it was to prove that it was Osama that had been plugged, and then they conveniently threw away the actual evidence for which we only have there word (until the videos are released) that existed in the first place.

    I would be happy to believe that the world is rid of Osama, but I do have to exercize all the usual scepticism given the amount of bias that could potentially be brought to the experiment. Peer reviewed science it is not.
    Oh, so now they have a mass spec sitting in Bagram now?

    Here is what New Scientist is saying
    "The agency responsible for conducting the analysis is also unconfirmed: media reports have named both FBI and the CIA. Several sources suggested that DNA was subpoenaed from the body of a half-sister of bin Laden, when she reportedly died at Massachusetts General Hospital in Boston, but the hospital was unable to confirm the report. "There is no evidence she was even here," says spokesperson Sue McGreevey.

    FBI special agent Greg Comcowich would not confirm or deny any investigative steps the FBI has taken. When asked whether the agency would release any further information on the DNA analysis, he replied: "Not that I'm aware of.""

    Of course, given the fuss Americans made about wikileaks, I assume if anyone did leak details of where and how this alleged genotyping took place, Fox News would be calling for that person's assassination.

    I repeat, I don't doubt that Osama really has been dumped in the ocean - I just don't believe a DNA genotyping has been done and this point to an increasing culture of contempt towards normal standards of open and honest government

    Well, a mass spec isn't any more or less likely than a capillary electrophoresis instrument. In fact, I wouldn't be at all surprised if they have both types of instruments, and then some, available in the area. Both are used extensively in forensic investigations of various types and if you don't think there's any battlefield forensics taking place over there, you're crazy.

    So if they did kill someone they thought was him, I see no reason why they wouldn't be able and eager to analyze the DNA to confirm it.

    Of course, as mentioned we'll never know for sure anyway. Photos, lab results, etc. can all be doctored/manufactured, so even a presentation of those things won't be enough for the truly skeptical/conspiratorial. Heck, there's still people that believe president Obama wasn't born in the US despite all evidence provided, so you could probably dump bin Laden's body on some people's living room couch and they still wouldn't believe it.

    Gerhard Adam
    The trouble is that most people can't distinguish between being skeptical versus simply being stupid.  If they spent a fraction of the time actually paying attention to the political process and using their votes properly, they wouldn't have a government that they despise.

    People never get the government they want, but they invariably get the one they deserve.
    Mundus vult decipi