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Okay... now with my trusty PocketPCR... I shall... what does a PCR do again?

by WrtCdEvrydy 6y ago

zitterbewegung 6y ago
It helps you sequence a genome . On the website they give examples like sequencing beer yeast.
- monocasa 6y ago
  Well, it replicates DNA. There's other uses for it other than just sequencing, but that is a common one.
- vikramkr 6y ago
  It doesn't sequence, it amplifies DNA/RNA (through reverse transcription) and lets you detect a particular sequence or produce more of a sequence you want to use for downstream steps
  - fastaguy88 6y ago
    No reverse transcription. Just amplification by extending a primer using DNA polymerase (rtPCR -- reverse transcription PCR -- of RNA does do an initial reverse transcriptase step.) The PCR process is also used in modern DNA sequencing, but that is not what a PCR machine does. And a non-qPCR machine does not detect the sequence -- it just produces enough of it by amplification so that it can be detected using other methods.
    - vikramkr 6y ago
      No, you can use a thermocycler for reverse transcription. In fact, you need one to do rtPCR, and I don't see why this cycler wouldn't be able to do that. And to clarify, I didn't say the device detects a sequence. I said it lets you detect the presence of a sequence. This isn't a biology forum, so I deliberately didn't go into excessive detail and technicalities, just conveying the information that one of the things you use this for is detecting a DNA sequence (and sure, there are post processing steps like running a gel or sending it off for sanger sequencing etc), and the other sort of thing you can do is RT-PCR. You can also do all sorts of cloning techniques like gibsons. And technically, neither device nor a qPCR machine let you detect a particular sequence exactly. It just amplifies a piece of dna enclosed by the relevant primers that you are using to amplify. I'm well aware of all these technicalities with saying that a thermocycler does sequence detection, I left them out on purpose. And as mentioned before, the pocketPCR, like any other thermocycler, is absolutely capable of doing RT etc. I know the parent comment said "what does a PCR do again?," but it's pretty clear from context and use of the article "a" that they are asking about the pocketPCR machine's capabilities, not about polymerase chain reaction in general. If they were, then yes you are technically correct regarding RT.
      - robbiep 6y ago
        This is overly pedantic but you did literally say lets you detect the sequence in a context where the average reader would have parsed the sentence as referring to the capabilities of the device.
joshuamcginnis 6y ago
A thermocycler is simple: it heats up and cools down in series of programmable cycles. The thing being heated and cooled is often a tube of DNA and reagents. The strands of DNA denature, or separate into individual strands, at a specific temperature and the reverse happens (annealing) at cooler temps. Imagine peeling apart a double strand of DNA creating two single strands. Now imagine making each single strand whole again by plugging in free-floating nucleotides. You'd now have two strands of DNA when you started with one. This is a simplified explanation of DNA amplification and is quite useful for taking a small sample of DNA and making lots of copies for analysis like sequencing.
vikramkr 6y ago
so your pocket PCR is a thermocycler, which is a thing that heats and cools samples of liquid to set temperatures for set times for a programmed number of cycles. Tube gets hot, tube gets cold, makes a few stops along the way, repeat. Now, what heating and cooling the liquid to precise temperatures a bunch of times actually does depends on what's in the liquid. The most common use case is something called Polymerase Chain Reaction (or PCR, hence the name of the product). As another comment explains, this basically lets you take a little bit of DNA and make a lot of DNA. You can run other reactions as well, such as reverse transcription, where you take some RNA and generate a DNA sequence out of it. There are a lot of reasons you might want to do that, but for example that's a common procedure in diagnostics when testing for viruses like SARS-COV-2. You can do molecular cloning work, which is editing and modifying DNA, stitching DNA sequences together, etc. It's a very useful took and a core component of most biological labs. Unfortunately, if you buy a PocketPCR, you end up with a device that doesn't have a heated lid. This means that you have to add mineral oil to all of your samples before cycling them since otherwise you could screw up the reaction/lose your sample. As someone who has dealt with that before, take my word for it that it's a royal pain. Splurge a bit on the heated lid.
numpad0 6y ago
Sounds like it heats up and cool down your vials in certain way, that allow enzymes sold separately to replicate DNA chunks of your specification, through Polymerase Chain Reaction reaction, so that DNA in question can be optically or electro-mechanically detected/observed by sheer volume of it

Okay... now with my trusty PocketPCR... I shall... what does a PCR do again?

zitterbewegung 6y ago

It helps you sequence a genome . On the website they give examples like sequencing beer yeast.

monocasa 6y ago
Well, it replicates DNA. There's other uses for it other than just sequencing, but that is a common one.
vikramkr 6y ago
It doesn't sequence, it amplifies DNA/RNA (through reverse transcription) and lets you detect a particular sequence or produce more of a sequence you want to use for downstream steps
- fastaguy88 6y ago
  No reverse transcription. Just amplification by extending a primer using DNA polymerase (rtPCR -- reverse transcription PCR -- of RNA does do an initial reverse transcriptase step.) The PCR process is also used in modern DNA sequencing, but that is not what a PCR machine does. And a non-qPCR machine does not detect the sequence -- it just produces enough of it by amplification so that it can be detected using other methods.
  - vikramkr 6y ago
    No, you can use a thermocycler for reverse transcription. In fact, you need one to do rtPCR, and I don't see why this cycler wouldn't be able to do that. And to clarify, I didn't say the device detects a sequence. I said it lets you detect the presence of a sequence. This isn't a biology forum, so I deliberately didn't go into excessive detail and technicalities, just conveying the information that one of the things you use this for is detecting a DNA sequence (and sure, there are post processing steps like running a gel or sending it off for sanger sequencing etc), and the other sort of thing you can do is RT-PCR. You can also do all sorts of cloning techniques like gibsons. And technically, neither device nor a qPCR machine let you detect a particular sequence exactly. It just amplifies a piece of dna enclosed by the relevant primers that you are using to amplify. I'm well aware of all these technicalities with saying that a thermocycler does sequence detection, I left them out on purpose. And as mentioned before, the pocketPCR, like any other thermocycler, is absolutely capable of doing RT etc. I know the parent comment said "what does a PCR do again?," but it's pretty clear from context and use of the article "a" that they are asking about the pocketPCR machine's capabilities, not about polymerase chain reaction in general. If they were, then yes you are technically correct regarding RT.
    - robbiep 6y ago
      This is overly pedantic but you did literally say lets you detect the sequence in a context where the average reader would have parsed the sentence as referring to the capabilities of the device.

joshuamcginnis 6y ago

A thermocycler is simple: it heats up and cools down in series of programmable cycles. The thing being heated and cooled is often a tube of DNA and reagents. The strands of DNA denature, or separate into individual strands, at a specific temperature and the reverse happens (annealing) at cooler temps. Imagine peeling apart a double strand of DNA creating two single strands. Now imagine making each single strand whole again by plugging in free-floating nucleotides. You'd now have two strands of DNA when you started with one. This is a simplified explanation of DNA amplification and is quite useful for taking a small sample of DNA and making lots of copies for analysis like sequencing.

vikramkr 6y ago

so your pocket PCR is a thermocycler, which is a thing that heats and cools samples of liquid to set temperatures for set times for a programmed number of cycles. Tube gets hot, tube gets cold, makes a few stops along the way, repeat. Now, what heating and cooling the liquid to precise temperatures a bunch of times actually does depends on what's in the liquid. The most common use case is something called Polymerase Chain Reaction (or PCR, hence the name of the product). As another comment explains, this basically lets you take a little bit of DNA and make a lot of DNA. You can run other reactions as well, such as reverse transcription, where you take some RNA and generate a DNA sequence out of it. There are a lot of reasons you might want to do that, but for example that's a common procedure in diagnostics when testing for viruses like SARS-COV-2. You can do molecular cloning work, which is editing and modifying DNA, stitching DNA sequences together, etc. It's a very useful took and a core component of most biological labs. Unfortunately, if you buy a PocketPCR, you end up with a device that doesn't have a heated lid. This means that you have to add mineral oil to all of your samples before cycling them since otherwise you could screw up the reaction/lose your sample. As someone who has dealt with that before, take my word for it that it's a royal pain. Splurge a bit on the heated lid.

numpad0 6y ago

Sounds like it heats up and cool down your vials in certain way, that allow enzymes sold separately to replicate DNA chunks of your specification, through Polymerase Chain Reaction reaction, so that DNA in question can be optically or electro-mechanically detected/observed by sheer volume of it