Polymerase chain reaction (PCR)
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- Опубликовано: 12 сен 2024
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Got a big interview tomorrow I'm pretty nervous about, this helped a ton. Not only was it informative, but it was also pretty calming. Thank you.
Did u get the job
Did you get the job?
@@prestongriffith4640 @radcow I got the job! Thank you for asking
@@MisterTwiin Congratulations
@@MisterTwiin 👏
straight to the point no bullshit. Need more people like this in the world
Please amplify ur sound as well ,it seems someone is whispering
Yes. I have to off my fan for hearing this.
3:15 he listened to you
Nice tutorial!, just for the record, it is thermus aquaticus (with a 'u')
Thanks lol, you just saved my ass
number of dsDNA molecules is 2^n, where n=number of cycles of PCR completed
More specific is amplicons = A*2^(N-2) where A is the number of template strands you start off with and N is number of cycles. You actually don't make any amplicons in the first two rounds.
Why can't we just add helicase to denature the strands rather than heating?
+Devika Satija I think because we can't shut them off when they need to be off. You see PCR requires three stages, and DNA double strands only need to be separated at stage 1 (denaturation), if we use helicase, it will be active at stage 2 (annealing - primers attach to parental strand) and 3 (elongation - polymerases add nucleotides) also, and we don't want that.
1) with heat we can separate entire strands at once, instead of just small local areas with helicase
2) it's much quicker and easier and efficient to use heat
It is difficult to maintain optimum conditions to use Helicase outside the living cells. So we use heat rather than enzyme.
thank you maybe a bit longer and more explicative but ok for a recap
Don't you need 2 primers? One for the sense and one for the antisense strand?
This video helped so much!
Hi, thank you so much for the video. Can you please tell me what exactly primers are and why we use them? I didn't get that really well.
Basically it’s an rna molecule, the polymerase starts building the complementary strand from that rna molecule. In the 5’ to 3’ the polymerase only require one such piece to be placed but on the 3’ to 5’ one it needs multiple and then it connects each two in 5’ to 3’ directions. When the process is done those rna molecules are replace with dna molecules and you get a double strand dna
In simpler terms, primers are added so RNA/DNA polymerase can make new daughter strands faster, think of them as checkpoints to where to start the new sequence. There will always be a lagging and leading strand,( in the case of PCR there are only leading strands(idk why there just is). The lagging strand has multiple primers while the leading strand will have way less because it's just a straight line. Primers are added in replication bubbles too. I can go on forever and example further but then I would be writing an essay.
ill try to draw a picture,
< = primer
---- = new strands
It's specific to the segment you want to copy. You use the primer that corresponds to the priming sequence on the DNA strand you want to replicate.
Intervention time! You say "Basically" at a faster rate than PCR creates DNA
were you recording with a toaster?
wow! clever AND original!
maybe with a hamster, who knows?
To
Thanks for the video. I've been trying to find out for a while why you would want to amplify DNA. Is it because endogenous DNA concentration/abundance is much too low for detection by current instruments? How does an instrument count the number of copies that are made? Many thanks.
InspirationIsFree Not an expert, but you'd most likely do electrophoresis from the DNA sample after having done a PCR so I suppose it's got to do with the original amount of copies being insufficient
+TheInsaneFellow We do not directly "count" the DNA one by one. According to what I've been taught, we do electrophoresis, like TheInsaneFellow have said, with ladders of known DNA concentration. Then we basically compare the width of the DNA strands to the width of the ladders to pick out the most closed number.
PCR:
denature the strand through heat.
Thermus aquaticus DNA polymerize - copy of the DNA.
add primer, starting point, nucleotides.
exponential increase in number.
the detail is so underwhelming
I mean, its a free non profit online course, were you expecting a Harvard lecture?
it's perfect for a quick introduction, but if you thought this was going deep in an under five minutes video, i want to know what other vids you use to base this on on.
M
Thank you great vid keep up the good work
Thanks that was rewlly helpful. .
*really
wow.
Leighton Mayers mmmm k
wow.
@@rashidamohammed8463 *ok
This is a little sloppy for the standards of khan academy. It's still good though
Helpful..!!!😀
Great video...your voice is a little too soft, difficult to hear. But the video helped heaps..Thank you!
Check the math when amplifying your DNA: it goes from 1 to 2 to 4 to ... 16! (See comment from Nick George below: the number of dsDNA molecules is 2^number of cycles of PCR completed.)
Check your own math.
2^1 = 2 (after first round)
2^2 = 4 (after second round)
2^3 (2*2=4*2) = 8 (after third round)
2^4 = 16 (after fourth round)
I hardly hear you.
thanks!
Where does taq source the nucleotides to build the complimentary strand? Would you need to add it in to be able to have thousands of copies?
+hunglikehuang There are free nucleotides in the solution (that's added). TAQ is just a DNA polymerase, which basically reads the strand of single stranded DNA and adds the correct complementary nucleotide to that single strand, resulting in a double strand now
+Nik Friddle Thank you
yes, you must add the dNTPs (deoxynucleotide triphosphates)
just a quick clarification, so by the end of the PCR cycle will always end up with a double stranded DNA?. in other words am asking, when going for Capillary electrophoresis, we go in with a double stranded DNA from PCR? thnks
Thank you ❤️
Sound system is not optimal please try to improve.
How does pcr stop dna synthesis?
What if there was only 1 primer, not 2 primers. What would the final product be?
My guess is that the DNA will not be amplified at all. Because if only one primer is attached to one of the single strand, only one of the 2 single strand will be elongated. As a result you will only get back just one copy of the same DNA,which defeats the purpose of PCR.
what did you mean by annealing?
Annealing is another term for attaching primers. It does the same job as primase which attaches primers. For PCR instead of using the enzyme Primase you Aneal it which means attaching primers, as i stated before.
thanks this helped 👍
If the nucelotides were removed, would you still get elongation of the DNA strands?
No, you won't
That's like building a house without bricks (or any other building materials)
Damn you. I need you to fly tomorrow
Video is too quiet. Can't hear anything
Turn up your 🔊
And that my friends is how you can get a positive test for corona...taadaaah!
🤣🤣
@Aguilar Denerde Wow! You're a real tool.
@Aguilar Denerde what are u trying to get at LOL
Looks like somebody's mad his daddy got booted off the big screen. And I've heard it's actually amplified 33 times. And Aguliar what you know about Karys collections?
@Aguilar Denerde honestly man what are you trying to say in your first comment
Is there any videos with RT-PCR?
Voice is low and talk so fast
he sounds like Kermit the frog
What possible complications can occur in PCR that would bring unwanted results?
The Taq polymerase doesn't have a proper proofreading mechanism - meaning it won't detect any wrongly placed nucleotides. The whole system is also subject to contamination between nucleic acids.
Can you explain to me how this test is able to detect a specific strain? Like.. covid19?
it can't bro ! COVID-19 is fake
Xander Smith I agree lol. I started asking these questions on all these scientific videos explaining these Mickey Mouse tests. No answers so far...
thatstheguy07 bruh google it
Billy the Goat bruh, I don’t need to.
thatstheguy07 if u need an explanation then yea you do, but it is up to you and I respect your decision.
So you show two strands and then jump to amplifying one of them without showing how we one was chosen 👎🏼 #fail
bad sound, sorry.
16 after round 3 not 8
JKDMan2000 no it's 8
JDK youre a dumb fuck
PAS OUF EN SAH
Once you realized that you had fucked up and explained things in the wrong order, you should have done the video over.
Underline your damn genus names!
Improve your sound quality
RUclips! Please tell me what the "late stage of PCR" is PLLEASE or else I'll diiieee
@Nyanna Ross don't die.. the last stage is final hold
Your audio recording is simply awful.