polymerase chain reaction

[jellyfishlollypop]

Thank you soo much, this was extermly helpful

[ladybugflv]

Thank you for answering at such a short notice!

[ladybugflv]

Quite an exhaustive video indeed. Still, what I would like to know is if magnesium ions are also needed for the Taq to build up the chains, as in any replication process. Moreover, I am also curious if the pH plays a certain role. I guess it needs to be around 7 or 8 and it has to remain constant, but I'm not very sure.

[gkpeter]

@ladybugflv Magnesium concentration is very important in this process. When troubleshooting reactions, it is one of the components you mess with. It is a cofactor for Taq and also helps stabilize dsDNA. pH is important in PCR as it is in any biological reaction. Drastic changes in pH can destroy the enzyme and also hinder annealing by preventing hydrogen bonding from occuring.

[proteinP53]

very good

[Enorim]

Thanks. As a rookie MB graduate student , these are just what I need. Great , explicit.Looking forward to your videos on Primer Design !

[isena]

hello, very nice video and helpful explanations. but my question is: why do we get only in the 3d cycle the product we need? thank you

[gkpeter]

It is not until after the 3 rd cycle you get a double starnded versionof your desired product. After the second round you get a single stranded version, then that will be used in the 3rd cycle as a template to create a double stranded version.

[isena]

ow, yes of course. but why is that? btw: thank you for the quick reply!

[gkpeter]

Well, it is just what happens. The first cycle will create a hybrid between the template and a fragment started with the primer. That fragment started from the primer will keep going until the polymerase drops off or the elongation cycle is over. What you should notice is that fragment is NOT the size you want. But what is important is that one end is started with your primer. So, in the next cycle the other primer will anneal and then drop off when it reaches the end created by the primer

[stephanies007]

Thank you...this video is very helpful.

[gkpeter]

No problem, if you need anything else explained, feel free to ask.

[DeMarth]

Nice explanation, thanks

[gkpeter]

No problem, if you need anything else explained, feel free to ask.