They can find the sequence of each small DNA fragment attached to the PoliA strand but how do they join all those fragment readings in the correct order to build the correct entire original genetic sequence?
I see that this method allows to sequence single molecules withouth previous amplification. How is it posible? I mean, other techniques such as Illumina or SOLiD require having several copies in order to produce a signal big enough. Why is it different here?
Yeah, that's what I thought, I was just making sure I heard things right. It's amazing how genetic engineering is performed these days. ^.^ With the information given by this process, we could eventually compare every known organism with each other and have the best map of evolution ever made (not to mention what this would do for other important fields, particularly medicine, but that's what I'm interested in).
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are there limits to reconstructing the genome due the 100bp size limit (due to vntrs, transposons)? also, if the average area per polyT binding site is 1 square micron, how accurate can the imaging be?
"This was a triumph, I'm making a note here.. huge success. It's hard to overstate my satisfaction" Doesn't the one narrator sound like the person from portal? Robotic at the least.
these third gen sequencing platforms are just revoluntionary but imagine the terabytes of data produced! thats where computation comes in. This is great
there are bioinformatic approaches called de novo assembly by Bujrin graphs. I can't be so concise to explain it in a youtube comment but if you read into it you read into de novo assembly you'll find answers. Also read on programs like velveth, ABYss, TopHot, Bowtie etc.
@montelka unincorporated nucleotides with the fluorescent marker in spots not tagged as targets would not be such a problem right? Software simply ignores them. Heck with a billion per run you can ignore some suspect signals too. Right?
Because of different method of gaining that signal. Here, the signal is gotten by the fluorescent addition being referenced by the fixed position of one molecule using an ultra high resolution camera. In Illumina, the signal is gotten through a pH change - one H+ will create such a small change that the unit will simply not be able to ID it, hence needing amplification.
interesante la plataforma helicos de secuenciamiento, because i really think that this method can be improving in the future to have a higher throughput
@pedrolikes2008 They are aligned by computer to a reference genome. So you cannot sequence something new, you rather look for differences in genome of something that was sequenced before.
I would assume that a computer program could be designed to look for overlapping sequences between pairs of the individual 100bp strands in its database. 100bp sequences seem long enough to ensure that the overlaps really constitute a true match, not just a chance pairing. I don't really know that much about programs like this, but it seems to me the the computer could be designed so that it knows the amount of overlap which is statistically significant.
Beautiful... after every wash of adenine, guanine, cytosine, or thymine they see what stuck, and in the end have each piece of DNA sequenced properly... just, beautiful... So an entire genome could be done in a day by one person? Biologists could have every animal mapped in no time... BTW: there is an old article on How Stuff Works about this, but it isn't as good as this vid IMO as far as describing the whole process.
Computers! You can use scripts to align the short sequences to each other - you have so much coverage of the genome that each location in the genome is present on many different fragments of DNA. "Stack" these to assemble the longer sequence: ________ ____________ ____ _________ ^^^ we can be pretty sure of this area. Imagine other fragments filling the gaps too!
@pedrolikes2008 After you do the chopping some fragments contain some part of the sequence of other fragments. A computer program puts the pieces together.
"So an entire genome could be done in a day by one person? Biologists could have every animal mapped in no time..." No, what they said was: "Sample PREPERATION is simple, and can be performed by a single person in less than a day." They did not include how long all the other things would take.
See my response to vitr1ol above... Any inaccuracies will hopefully sufficiently "covered" that we can call it a random error too, but the imaging is rather precise. It knows where the laser is pointing at any moment and the detection limit is quite low.
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