Talk:Polymerase chain reaction

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Old comments

There appears a concept called "Tm" in this article, but it is neither linked to an article with such title, or explained in this article itself. Please, define this term in its own article (and link to it), or define it here. Thank you. Pentalis 21 april 2:01 GMT -4:00 winter time.

• It stands for melting temperature, often abbreviated Tm in calculations. Not sure why it is Tm rather than Mt, though capital T usually stands for temperature. CheekyMonkey 12:49, 2 May 2005 (UTC)

• • Thank you very much, I also noticed you added the concept to the disambiguation page too, very kind of your part :-) Pentalis 6:25, 31 May 2005 (GMT -4:00 winter time)

Fixed to T_m. 203.218.136.155 14:39, 11 June 2006 (UTC)

In step 4 of figure 2 is not explained. How is the rest of the DNA past the primers truncated? All we see is a dotted line on the diagram suggesting it is cut somehow.

They don't get replicated, and hence drop out of significance entirely. (Among the millions of DNA molecules that eventually get made.) Its just a diagram to illustrate the concept, the actual reason is in the article. -- Natalinasmpf 13:27, 16 Jun 2005 (UTC)

CATTAGAT

There's a new program being released (under the GNU GPL) to perform primer specificity searches.

http://genome.cs.hi.is/cattagat/

If this looks useful to anyone then perhaps it can be added to the external links. I don't feel right doing it myself since I'm one of the authors of the software. - Haukurth 15:55, 21 July 2005 (UTC)

Har Gobind Khorana

I don't see how one can attribute the invention of PCR to Khorana. An invention (at least a patentable one) requires a reduction to practice, which he certainly never did. There is no reference cited for this either. In addition, the theromostabile polymerase was not known to Mullis at the time of his invention. Mullis got the Nobel Prize for the invention, not Khorana. I am therefore removing the attribution to Khorana.DonSiano 17:23, 22 August 2005 (UTC)

• Hi DonSiano. I left the actual reference to the paper (which you removed), as well as the actual text from the paper. In the paper, (and in lots of subsequent work), they used the procedure (which I quoted from the original paper) to construct genes in their study tRNA related genes. Some of the other references mentioned in the citations at the bottom of the article also mention Khorana's work. The actual text you removed is below:

• Much of the conceptual work on polymerase chain reaction was carried out in Har Gobind Khorana's lab at MIT. Khorana and coworkers published a paper in 1971, describing repair replication and the process by which it could be used to synthetically replicate tRNA genes in E. coli. The major finding is given in this text:

The principles for extensive synthesis of the duplexed tRNA genes which emerge 
from the present work are the following. The DNA duplex would be denatured to 
form single strands. This denaturation step would be carried out in the presence of a 
sufficiently large excess of the two appropriate primers. Upon cooling, one would hope 
to obtain two structures, each containing the full length of the template strand 
appropriately complexed with the primer. DNA polymerase will be added to complete 
the process of repair replication. Two molecules of the original duplex should result. 
The whole cycle could be repeated, there being added every time a fresh dose of the 
enzyme. (Kleppe et al., 1971)

• I actually redundantly reference the work. If you do not access to the reference, I would be happy to send it to you. In fact, if you read the paper, they actually reduce it to practice not only in the referenced paper, but in much of the subsequent work in synthesizing tRNA related genes. Regardless, the wording I used was being conservative and not attributing an invection to Khorana's lab, just that the conceptual work was laid down over a decade earlier. The quote that I use from the paper (12 years before the invention of PCR) is PCR, they just call it something different. In fact, in the referenced paper, they determine other conditions that are conducive to such amplification, such as minimal primer sizes. I could reference Khorana's subsequent work on amplifying various pieces of genes using the procedure. Granted, Khorana's group did not push automation, thermostable polymerases, et cetera, but it is no reason not to mention and reference the work.

I'll post the information back, unless you can explain to me why this work is not important to the history of PCR. If the wording seems controversial, maybe we could consider rewriting the section?\

--Skosuri 03:46, 9 June 2006 (UTC)

---

Hi Skosuri,

Well, PCR is one of those really big inventions that someone other than the almost universally acknowledged inventor thought of first. And the history of the invention is always more complicated than that which can be accommodated in a paragraph or two of an encyclopaedic article. Are there any inventions that don't have that characteristic? Both the courts (and presumably the nobel prize committee) considered this dispute and came down on the side of Mullis. There is no evidence that Mullis was aware of Khorana's work; and others certainly contributed to making it work: including, Henry Erlich, Norman Arnheim, Randall Saiki, Glen Horn, Corey Levenson, Steven Scharf, Fred Faloona and Tom White. And maybe even Kleppe has a claim too. There is, understandably, quite an impulse to knock the hero down a bit. The LSD ref is attributable to this tendency too, I think. I note that the "history" section of the article makes no mention of the others either. Nor is there any note taken that Mullis was a synthetic chemist put out of work by a machine to synthesize oligimers...

The fact is that PCR could have been invented years before Mullis came along, and no doubt a lot of people regret they missed it. One has to wonder, though, how many years would have passed before it was developed if Mullis hadn't done his bit...

The bottom line is, I don't think Khorana deserves all the space you would like to devote to him it this article, which would seem to outweigh Mullis' contribution. I certainly don't dispute that some mention might be made, but it is out of balance. Perhaps you could try some rewording of this: perhaps "like most inventions there are precursors and other actors than the one..." or something along those lines. Please don't just revert--let's try to reach some middle ground that is acceptable to us both.

There has been a lot of interest in PCR as a prototypical invention and it has been the subject of quite a bit of historical work, as I am sure you are aware. Perhaps what is really needed is a separate article on this, perhaps "PCR History"; I'd be willing to work on it with you--it could be interesting and worth doing. We could move a good bit of the history section over there and leave a ref to "main article" here. Which could shorten the PCR article which is already too long besides. What do you think? DonSiano 12:29, 9 June 2006 (UTC)

Call it History of polymerase chain reaction if you do it, to be consistent with other articles 203.218.136.155 14:39, 11 June 2006 (UTC)

"PCR reaction"

Is it considered acceptable to use the phrase "PCR reaction" or is this considered redundant? The phrase appears a couple of times in this article. (RAS syndrome and the associated discussion page provide an interesting treatment of the issue; a Google search for "'PCR reaction' and 'RAS syndrome'" also reveals some interesting hits).--GregRM 15:17, 21 December 2005 (UTC)

I try and remove the 'reaction' from the 'PCR reaction' whenever I can as it bugs me but maybe I'm just picky! I think it stems from the fact that acronyms such as PCR, ATM, CCTV and so on have become so commonplace it is no longer considered necessary to define them and so people lose sight of the actual meaning and consider the concept as a whole. Thus, when wanting to emphasise the fact that it is a reaction, the 'reaction' tagged onto the end. -luckyluke09

Yes it is redundant, always remove reaction coz this is an encyclopedia 203.218.136.155 14:39, 11 June 2006 (UTC)

Thanks for the replies...I removed references to "PCR reaction" yesterday.--GregRM 22:35, 12 June 2006 (UTC)

"PCR reaction" is not only acceptable, but also the appropriate wording when the sentence context requires the word "reaction". Here is why: The term "PCR" is a name, i.e the name of a significant and widely known method, not a mere abbreviation or a shorthand for the syntagm "polymerase chain reaction". The latter is simply the term describing any technology that would utilize polymerase in one exponential process, as opposed to Polymerase Chain Reaction, which is one of the two accepted names (mutual aliases) of the known and invented process of an enormous significance. Were "PCR" only a written shorthand, then we would read it using all three full words, but we don't. As we see from this, the word reaction is neither repeated nor redundant. NenadZILIĆ (talk) 19:06, 6 December 2019 (UTC)

History of PCR

• History of PCR A great review article. Also PCR Protocols, PCR Bioinformatics, and discussion are included.

Links to other PCR topics removed as they are not informative for standard PCR, and should be under Real-time or quantitative PCR. The History of PCR link is much more informative for PCR, yet was removed by [user:Alan Au] -- Bioinformin

Taq is not a polymerase that is used always

I disagree strongly with the phrase that Taq is used in current practice. Taq is with current standards a bad polymerase and it is not necesary to use Taq. Taq has no proofreading activity and its processivity is not as good as other polymerases, so it is my opinion that wikipedia should take an independent stance and not link PCR to Taq

I also made a very imortant change. It is not nucleotides that are used in the reaction but deoxynucleotides-triphosphate

I think you mean deoxynucleoside-triphosphate. There is no such thing as a nucleotide-triphosphate, since the definition of a nucleotide is a nucleoside + phosphates. 130.243.248.239 14:29, 19 November 2006 (UTC)

Comment:

yes i completly agree to this. while taq polymerase certainly is the most famous and perhaps the one most used, it is far from the only one, nor is it one of the best polymerases available. an elucidation of the other available polymerases would be much appreciated. here assessing their properties, comparing price (relatively, not absolutely), areas of use. where it is appropriate to have a polymerase with high fidelity. 3' nucleases 5' nucleases. and so on and so forth.

thank you -broccolee

Bicycle or Car?

I was told that Mullis was driving his car and that he was going on vacation with his (first) wife and that he got the idea that he just had to try, so he turned the car around to the lab... Is the motorcycle source reliable?

Several theories abound, ranging from the drugs story to the possibility that Kary Mullis merely reduced to practice the idea that had been given to him by a co-worker who had attended a seminar discussing tRNA synthesis in a manner similar to what we now know as PCR. Another rumour is that Kary Mullis was not going to be named as an inventor on the patents (the glory instead passing to David Gelfand, a co-inventor on the Taq patents, and Randy Saiki), and came to work at Cetus one day with a gun, demanding to be named as the lead inventor. All rumour though, and not suitable for the main page. 159.92.30.11 12:53, 1 June 2006 (UTC)

elongation vs extension

i observe that one use the term elongation for the polymerase 'elongating' the primer along the dna strand. however the more correct term is extension. also elongation is used regarding protein synthesis.

the analogy is

melting-annealing-extension:PCR-cycle::intiation-elongation-termination:protein synthesis

i am fairly sure these terms are not used over one another. however you may know otherwise.

thats all.

micro-PCR

In my research on microelectromechanical systems (MEMS) devices, I ran across some information on micro-PCR devices, which are fabricated with MEMS techniques. They consist of arrays of reaction chambers with electric heaters, and their construction allows them to do faster PCR with smaller sample sizes (both DNA and reagents). Does anybody else know the history of micro-PCR (sometimes denoted μPCR) that can help me add some stuff on this topic? Good article: http://www.sciencedirect.com/science?_ob=MImg&_imagekey=B6TFC-4CPM20J-1-1&_cdi=5223&_user=1238599&_orig=search&_coverDate=01%2F15%2F2005&_qd=1&_sk=999799992&view=c&wchp=dGLbVlb-zSkWz&md5=ded7b4a9d79b6ef6bc883c7d08e00020&ie=/sdarticle.pdf

Could somebody please include Partial DNA match explanation in this article ?

I think partial DNA match is relevant topic to be included in this article. Could somebody please do that ? I need information about Partial DNA match interpretation. For example, it test is done on 13 alleles, and match is partial (on 10 of them), and reaction has not been succesful on 3 alleles: how should this be interpreted ? As far as I know, there are divided opinions in scientific community about this issue. Please help, this issue is important to me, and at the same time I think it is relevant to be included in this article, because it makes it more complete and understandable.

possible rephrasing needed

"Primers are short, artificial DNA strands — often not more than fifty and usually only 18 to 25 base pairs long nucleotides that are complementary to the beginning and end of the DNA fragment to be amplified." How can a nucleotide be 18-25 base pairs long, this needs to be rephrased.

removed nucleotide. 203.218.136.155 14:39, 11 June 2006 (UTC)

"Colony PCR - Bacterial clones (E.coli) can be screened for the correct ligation products. Selected colonies are picked with a sterile toothpick from an agarose plate and dabbed into the master mix or sterile water. Primers (and the master mix) are added - the PCR protocol has to be started with an extented time at 95^^C." Would you really use a sterile toothpick, Wouldn't a sterile wire loop be more appropriate? --KX36 10:47, 27 May 2006 (UTC)

ile code

Can someone explain why the DNA code for ile is AT(A/C/G) when the RNA code is AU(A/C/U)? I thought RNA is complementary to DNA so shouldn't the DNA be TA(A/G/T).

Explanation: If you recall, it is the antisense (noncoding) strand of the DNA that is used as the template strand for the mRNA, meaning that the actual coding DNA strand and the mRNA produced during transcription will be identical except for the G->U nucleotides. Here's a website with a few more diagrams to help: Transcription Amasa walker III 03:48, 23 June 2006 (UTC)

Redirection

Primer extension is not the PCR reaction! It's a method used in 5'termini mapping!

nomenclature

Replaced several descriptions "ionic" bonding with hydrogen bonding. Made some other changes. Am very new at Wikipedia editing but have been doing PCR and related technologies for many years. User:Twbeals31 26 July 2006

Is it a good example of PCR elecrtrophoregram

Why to put such a bad electrophoresis gel photo in the article? It is quite noisy and really bad. There are billions around for example mine. Does this have a some significance? --Araks 13:36, 16 September 2006 (UTC)

I agree. I work in a lab as well, so I can provide a better image.--Roland Deschain 18:41, 16 September 2006 (UTC)

Ok. I've added a gel from my own research. It's reasonably clean and produces one specific band for each reaction.--Roland Deschain 19:06, 16 September 2006 (UTC)

Use in identification of pathogens

There are several infectious diseases which require the use of PCR for identification of the causative agent as alternative techniques require incubation periods which render them useless. shouldnt this be added to uses?

GA Re-Review and In-line citations

Members of the Wikipedia:WikiProject Good articles are in the process of doing a re-review of current Good Article listings to ensure compliance with the standards of the Good Article Criteria. (Discussion of the changes and re-review can be found here). A significant change to the GA criteria is the mandatory use of some sort of in-line citation (In accordance to WP:CITE) to be used in order for an article to pass the verification and reference criteria. Currently this article does not include in-line citations. It is recommended that the article's editors take a look at the inclusion of in-line citations as well as how the article stacks up against the rest of the Good Article criteria. GA reviewers will give you at least a week's time from the date of this notice to work on the in-line citations before doing a full re-review and deciding if the article still merits being considered a Good Article or would need to be de-listed. If you have any questions, please don't hesitate to contact us on the Good Article project talk page or you may contact me personally. On behalf of the Good Articles Project, I want to thank you for all the time and effort that you have put into working on this article and improving the overall quality of the Wikipedia project. LuciferMorgan 02:17, 16 December 2006 (UTC)

Article is too big!

I propose moving the primer design information to Primer (molecular biology). I also propose creating a new page PCR optimization and moving all the PCR optimization information there as well. -Madeleine 00:46, 23 March 2007 (UTC)

first image is not definitive

the first image that's pcr tubes are not typical image for pcr. I am molecular microbiologist and never saw such colorful pcr mix. appealing image but not suitable I think. ArazZeynili 11:23, 21 April 2007 (UTC)

Neither have I--taqman probes have a red hue though, so it's possible that it's a fluorophore-based PCR. There's apparently some precipitate at the bottom of the tubes, lending credence to the fact that it's a colony PCR (which may have caused the coloration) as per the legend. It would be best to track the editor who inserted the pic to get more info. Malljaja 13:21, 21 April 2007 (UTC)

I had the same thought when I saw it added, but a quick googling for "colony pcr" found me this protocol which uses sucrose red in the mixture: http://www.cbs.umn.edu/~kclark/protocols/bactPCR.html. I've seen this before, in pre-made form -- dyes and some heavier stuff is added to the PCR mixture, this allows people to directly load the PCR solution onto a gel after the reaction is done, without taking an extra step to mix in loading buffer. (I didn't personally use it, someone else in the lab had bought one of these products.) Anyway, here's some examples of pre-made versions: [2], [3], [4].

Although the image wasn't typical, it wasn't technically wrong either. Not wanting to look a gift horse in the mouth, I didn't take issue with it at the time. One of us could certainly take a pic of a strip of PCR reactions next time we set one up and replace this pic, if you think the color is going to throw people off. I wonder if there should be a hand in it, for scale. -- Madeleine 14:40, 21 April 2007 (UTC)

I would put this 'gift' down in the article. person facing pcr for the first time will think that this is very typical pcr thing which is not. hand for scale goog idea ArazZeynili 22:52, 21 April 2007 (UTC)

I've replaced the picture with one of my own, a strip of eight 100ul PCR reactions I set up. Madeleine 15:55, 14 May 2007 (UTC)

Restructuring article

I think the history should go higher, but after the basic description of PCR. What do people think about rearranging the article into this form:

• The polymerase chain reaction (description of basic protocol/process)

• sample procedure

• History of PCR

• patent wars

• Applications of PCR
• PCR optimization and variations (many modifications, eg hot-start PCR, have been developed as optimizations, so I think the two subsections belong together)

• optimization
• modifications to the technique

I'm open to different orderings, of course. Mainly I'd like to move the optimization and variations away from the core "this is how PCR works" section. Madeleine 19:08, 19 May 2007 (UTC)

Hi Madeleine, I do not feel very strongly about any re-arrangements. To me it seems that the practical parts almost deserve their own articles with "PCR" covering the basic principle, application, and history. The major shortcomings of the article were convoluted and redundant text and misconceptions creating mystification, which thankfully have largely disappeared, but may crop up again every now and then. I'd suggest you go on with any changes you feel are necessary--I'll offer any suggestions if necessary and time permitting. Many thanks! Malljaja 19:07, 20 May 2007 (UTC)

Gene clonning

The gene clonning section in this article is of no relevance, as far as I can see. PCR can indeed be used for detection of genetic modifications, but anyways, the entry does'nt seem to spell out anything of that sort. ώЇЌĩ Ѕαи Яоzε ^†αLҝ 23:46, 11 June 2007 (UTC)

Using PCR for gene cloning (or cloning of any piece of DNA for that matter) is one of the most common methods used in a molecular biology lab nowadays; so it's extremely relevant and really belongs here!!! To improve the section on PCR-based gene cloning, it should be expanded to include methods such as TA-cloning, overlap-PCR ("PCR splicing"), linking to nested PCR, and highlighting of potential pitfalls (cloning of long DNA fragments, proofreading, etc, much of which is already alluded to in other sections). I'll try to get to that soon, but anyone familiar with the techniques is encouraged to chip in! Malljaja 08:57, 12 June 2007 (UTC)

I guess just because TA cloning protocol uses treminal transferase property of Taq we can't use it over here. Indeed Taq is an enzyme used in PCR but exploiting the property is just an ease of clonning PCR products directly which is gained by using special vectors rather than special PCR protocols.

TA clonning is a protocol used to clone genes, but its not a PCR procedure used in cloning per se. It is more to do with the property of Taq (and similar DNA polymerases) rather than the PCR protocol. If we start including everything that uses PCR or PCR products then we will end up with the largest entry on wikipedia. In the latter case we ill have to include DOT BLOTS, SUBSTRACTIVE HYBRIDISATION, and even TRANSCRIPTOMICS!!! ώЇЌĩ Ѕαи Яоzε ^†αLҝ 09:37, 12 June 2007 (UTC)

PCR is an extensively used technique permeating through molecular biology like water through cloth, and the length of the article simply reflects that--as long as the PCR principles and application are conveyed in a straightforward non-redundant prose (something that has improved only recently), I do not see any problem with it (the index at the top allows easy jumping to the relevant sections). Moreover, there's been discussion about moving sections out of the main article, which has happened, e.g. with PCR optimization, which is a good way to deal with methods that require more detailed description and deserve their own article. Nonetheless, PCR-based cloning needs to be in the main article, perhaps broken down to nuts and bolts (if given in more detail elsewhere). TA cloning is indisputably based on PCR (as you yourself pointed out), and should be dealt with as such.Malljaja 11:07, 12 June 2007 (UTC)

Sorry, let me not be considered rude. I think we will leave it for others to comment. My point is, clonning is not a PCR protocol. Moreover the article entry on clonning doesn't spell out anything about TA or the terminal transferase property of Taq. May be if it does we can decide on if it fits in or not. On a lighter note, I do TOPO quite regularly and I have started hating minipreps. ώЇЌĩ Ѕαи Яоzε ^†αLҝ 11:13, 12 June 2007 (UTC)

Thanks for clarifying--now I see what you're getting at. I completely agree with you that the section on PCR-cloning contains precious little about the properties of Taq and how they are deployed in PCR-cloning techniques, and I think those belong in there for completeness (see my earlier comment). So I'd be happy to see such info in this section--if need be, PCR cloning would be seeded as a new article, and only referenced in the main article. Yes, may be others have thoughts on that as well. TOPO gave me some headaches in the past, so I've been doing most of my cloning by PCR (incidentally!). Malljaja 12:48, 12 June 2007 (UTC)

Most of these "uses" of PCR listed here are not PCR protocols, it's not just the "cloning". Many of the methods are here for the same reason cloning is listed -- they greatly benefit on PCR's ability to isolate a particular genomic region from whole genomic material. (I think this cloning word is confusing, it seems to refer to recombinant DNA / transgenic usage, which does greatly depend on isolating a particular region, not so much about isolating clonal DNA sequences.) All of these techniques (listed here) could be done without PCR but now use PCR to rapidly isolate the desired region:

• genetic fingerprinting

• paternity testing

• (sequencing)

• detection of diseases
• genotyping of specific mutations

• "cloning" (gene isolation -> recombinant DNA)

I've clustered these, they look like they could be condensed, and all of them could go under a single subject eg, "PCR is widely used as a tool for isolating and amplifying a particular region from whole genomic sample. Although it is possible to isolate fragments without PCR, this ability of PCR has made some techniques much easier to perform and therefore enabled their widespread usage."

Another major usage of PCR would be amplification of very low copy number -- although there's non-PCR methods of amplification that have less bias (ie. RCA methods). This overlaps with forensic usage (DNA fingerprinting is doable without PCR, but PCR enables genetic analysis for many samples that wouldn't otherwise be analyzable). The following arguably depend on this ability:

• (viral DNA detection)

• AIDS testing

• analysis of ancient DNA

The only remaining "use" here is the "comparison of gene expression" using quantitative PCR. Of course, we could always just hybridize unamplified cDNA to a microarray and never use PCR, right?

I think the entire section could needs to be reorganized, possibly under these major uses for PCR -- "isolation of genomic regions", "amplification of small amounts of DNA", "quantitative PCR". What do people think about this? In the meantime, I'm off to do some colony PCRs off my TOPO TA cloned bacteria.  ;-) -- Madeleine 17:35, 12 June 2007 (UTC)

I reckon Applications of PCR should be one single section. Types of PCR protcols should be expanded extensively either here, or seperate entries. Moreover as I said earlier I dont see why entries were added without citations. Anyways, I think this article on the whole needs some copy editing. If there are others who feel the same, please express your views. Because many students do refer to wikipedia for basic knowledge on things like PCR and it needs to be clear, to the point and with proper citations. Cheers ώЇЌĩ Ѕαи Яоzε ^†αLҝ 20:40, 12 June 2007 (UTC)

I can't tell whether you're agreeing with my proposal or not. -- Madeleine 20:54, 12 June 2007 (UTC)

I like Madprime's suggestions of re-organizing the article where necessary--I think she's already done a great job in recent edits (the article has been in much worse shape just a couple of months ago and has much improved since), so I trust that any changes she'll deem necessary will be sound ones. Wikiality you're preaching to the converted, I also think the article is lacking enough citations, and this small number of citations was a major reason for a recent downgrade in a Wiki review. Rather than lamenting the lack of citations, however, energy might be better spend finding and inserting references, something you're more than welcome to do (it's on my to-do list as well). Malljaja 21:32, 12 June 2007 (UTC)

Sorry Malljaja, I think this is getting an unfortunate personal U turn. I will exit here if thats the case. Let me make it clear. PCR is not a clonning protocol. Madeleine agrees to that too. I bumped into this article last night when I was editing (including adding reference) to GC-content article. You can check the article for the changes I have made. Anyways, my point is, if no one is fighting, that the applications should be in one single section, and doesn't need much explanation for each of them. I think we can all agree on one point, that PCR can be used as a first step for loads of applications. Just because PCR is used as the first step leading to something that can't be called as a seperate PCR protocol, same as isolation of DNA. Just because DNA needs to be isolated as a first step that doesn't mean that all the applications would be explained in details under Isolation of DNA. If we agree on this it would be nice. I appreciate the efforts taken by Mandeline and all others in fixing the article, but I think we still need citations for many things stated here. Anyways, I write in wikipedia to share my years of research experience, but not to be criticised for preaching. Sorry if I was anoying, I think I will leave this talk page to save my own dignity. Thanks ώЇЌĩ Ѕαи Яоzε ^†αLҝ 21:45, 12 June 2007 (UTC)

Just to add one more thing, citation is requested not because others cant find it, but because it is easier for the ones who wrote it to cite, because after all they would have read it somewhere. ώЇЌĩ Ѕαи Яоzε ^†αLҝ 21:52, 12 June 2007 (UTC)

Changes have been made

I've rearranged things quite a bit. Some references have been added, in the process, but please appreciate that adding references for already existing material is pretty tedious.

The list of PCR variations has been moved to the bottom, since it's pretty long and lacks narrative order. It may be possible to cluster it a bit. Some are PCR optimization, meant to reduce amplification of incorrect product (Hot-start PCR, Nested PCR, touchdown PCR). Methylation-specific PCR is related to allele-specific PCR (it's basically an allele-specific PCR where the "allele" is a uracil vs. methylcytosine). Both inverse PCR and TAIL-PCR are used to isolate unknown sequence next to known sequence (as I understand it)... but TAIL-PCR is also somewhat like nested PCR... -- Madeleine 04:53, 13 June 2007 (UTC)

Hi, I was reading the article and agree that the list of variations could be clustered under headings. I didn't give so much thought for a solution right now but I think hot-start and touchdown are a good grouping. Methylation-specific and allele-specific works. Inverse and Tail are good. I would list nested as an alternative to gel electrophoresis separation of PCR reaction products since that is all that is intending to do, but I could see how it is similar to touchdown also with the specific annealing and amplification, maybe list it under both. It occured to me that assembly and overlap extension are conceptually very similar and that the overlap extension PCR page could even be listed as an engineered form of assembly PCR, but perhaps that page's listing of ways to introduce different mutations in the PCR product compared to the template DNA merits its separate status...

I think maybe listing the broad goals of the modified PCR reactions would be a good way to group them but probably it would have to be a compromise between similarity in goal of modification vs. method/particulars of modification. Giftedlyarsenine (talk) —Preceding undated comment added 18:17, 16 June 2013 (UTC)

PCR Station

The premier site for researchers conducting PCR, helped me quite a bit during my Ph.D.

PCR —Preceding unsigned comment added by Testtubeboy (talk • contribs) 10:10, 2 July 2007 (UTC)

Going by Wikipedia's external links guidelines ... This site appears to aggregate paper abstracts and tacks on some advertising links. It is far from being "accessible to the reader", and it contains significant amounts of advertising. The claim that it helped you quite a bit during your PhD is actually an argument against its inclusion: Wikipedia is not a manual, guidebook, or textbook, there is a strong argument against providing or linking to content intended to aid use of PCR protocols in research.

The external links section of wikipedia pages often degenerates into a free-for-all advertising section for various links related to the subject of the page. Wikipedia is not an advertising space. For that reason I've tried to clean up the link section and trim it down to a core of links significantly useful additional content to readers. The PCR Station link looks like this sort of low-content advertising to me. In addition, I distrust your intentions in adding this link; you have made no contributions to wikipedia other than trying to add this link. Madeleine 15:11, 2 July 2007 (UTC)

LSD

• Better reference for Kary Mullis's LSD use

Is his own autobiography, Dancing Naked in the Mind Field. Here is a NYTimes review of the autobiography that mentions the LSD use and a site that has copied an excerpt from the book. http://www.csp.org/chrestomathy/dancing_naked.html http://www.nytimes.com/books/98/10/11/reviews/981011.11teresit.html Kevin143 08:47, 22 September 2007 (UTC)

Real-Time PCR

I suggest an addition to the "Variations on the basic PCR technique" section as follows:

RT-PCR: (Real-Time PCR) is a powerful and rapid technique for nucleic acid amplification. The accumulation of specific products in a reaction is monitored continuously during cycling. This is usually achieved by monitoring changes in fluorescence within the PCR tube. Touchstone42 13:50, 15 October 2007 (UTC)

Real-time PCR is already among the different techniques (under quantitative PCR) that appear under "Variations..." Have a look how it fits in there. If you talk about fluorescence, it needs to be specified from where the fluorescence originates. Also, its main use is quantification; your lead sentence basically describes PCR and doesn't mention quantification. Please note that RT-PCR is the common acronym for reverse-transcription PCR, so it would be inappropriate to use it here (qPCR or qRT-PCR (for quantitative reverse-transcription PCR) are the most commonly used abbreviations). Malljaja 15:31, 15 October 2007 (UTC)

I agree with everything you say. RT-PCR is the common acronym for reverse-transcription PCR however it is also a very common acronym for real-time PCR. I have found RT-PCR in many many places referring to real-time PCR. I think this needs to be accepted. Also, (I may be wrong) but quantitative PCR does not have to be real-time and real-time PCR does not have to be quantitative so I don't think the two terms are interchangeable. Touchstone42 15:31, 16 October 2007 (UTC)

I've likewise seen real-time PCR abbreviated as RT PCR in the literature. Some researchers have even dubbed reverse-transcription real-time PCR as RT-RT PCR, which sounds a wee bit like a stutter though. Ultimately, context is important, and acronyms often follow local conventions in the literature (i.e. full name followed by a, sometimes whimsical, acronym in parenthesis). However, as this entry gives a comprehensive overview of PCR and its different twigs and branches, simply calling real-time PCR "RT PCR" would be inappropriate, as it collides with "RT PCR" as used for reverse-transcription PCR. "Variations..." mentions the different naming conventions and conflicts, and it also touches on the issue that quantitative PCR methods are not always based on real-time PCR. Perhaps this section can be improved so that some of the naming and other issues can be put to bed. Malljaja 16:27, 16 October 2007 (UTC)

GA Review

This article does not meet the Good Article criteria at this time. Please see this page for the complete review. Dr. Cash 06:49, 23 October 2007 (UTC)

I've had a first stab at improving the article, mainly copy editing (i.e., boldly paring back unnecessary and convoluted wording), and inserting of refs as per the review. I've not done much work on the "History" section yet. This entry has slowly been improving, but it still needs come cleaning up--its previous length seemed to have invited some redundancy, manifested as overlinking and lack of reference to other sections, and some disorganization to creep in. Making it more succinct might help keep it more stable and to the point. Some of the the figures (Fig 3 in particular) look fairly poorly, and the text needs some more polishing to make it more accessible to the layman audience. I'm going to leave it for now, to give others the opportunity to chip in where they see fit. Malljaja 16:19, 23 October 2007 (UTC)

Too technical for the general reader

I agree that the article is too long, however I feel there's a bigger problem.

The article is much too technical for the average reader. There are far too many scientific terms. Reading the article I found myself quickly needing to either follow the links or grab a dictionary.

A wikipedia article shouldn't require additional reference materials in order for the general reader to gain a grasp of the basics of PCR (or any other topic for that matter; I could easily write an article on any number of Roman Emperors that would leave the general reader feeling just as out of depth as I do with this one).

In my opinion this article needs a rewrite to make it more accessible for readers not intimately familiar with molecular biology and related disciplines.

This article Kinetic energy penetrator on a ballistic technology does a much better job, again in my opinion, in explaining the subject without a blizzard of recondite terminology.

Therefore I've added the "cleanup-jargon" tag.

PainMan 08:40, 28 October 2007 (UTC)

I agree with you that there's a fair amount of specialized language in the article. It would help however if you were more specific--which sections are in your opinion most affected? I had a look with an eye on jargon, and the section that I see most plagued by it is the "Uses of PCR" section. Unfortunately the article you chose as a positive example (the KEP) lacks citations, raising the question of how much of the true complexity of the subject it reflects. Similar to other areas in the natural sciences--quantum mechanics come to mind (see e.g., Quantum computer, which if scaled to weather phenomena represents a class 5 hurricane in terms of arcane terminology)--articles dealing with molecular biology may not unlock themselves on first reading, and likely require some additional reading in other related entries. Sequence alignment is a featured article covering a fairly complex subject, so perhaps it could be the guiding light. Malljaja 12:17, 28 October 2007 (UTC)

PCR "Reaction"

Since the 'R' in PCR is for the word 'Reaction' its a tautology to say PCR Reaction. This means Polymerase Chain Reaction Reaction. I suggest the removal of all such occurrence within the text. —Preceding unsigned comment added by 129.215.149.97 (talk) 23:38, 9 March 2008 (UTC)

Agreed. You're most welcome to be bold and remove them yourself. Adrian J. Hunter^{(talk•contribs)} 02:14, 10 March 2008 (UTC)

I've gone through and changed the ones I found. Notably, sometimes "PCR reaction" is used to refer to the mixture in the tube and in some cases it I think it's preferable to say "PCR mixture" rather than simply saying "PCR" (which refers more to the process than the mixture in the tube). Madeleine ✉ ✍ 03:05, 10 March 2008 (UTC)

PCR machine pics

Wow, those are impressive! Very cool images of old machines. I think maybe they should go in the history section though. PaleWhaleGail also added a pic of a modern machine to the thermal cycler page, I think that would be more appropriate to the principle/procedure section. Madeleine ✉ ✍ 03:15, 14 March 2008 (UTC)

I'm glad you like the picture of the old machine. I've seen a few of them on the way to the skip. Young ones are shocked when they realise what it is. I wonder what they'd think of having to add kelnow fragment every cycle! If you want to put the three water bath machine in the history section, i think it might work well. Agesworth (talk) 08:14, 2 April 2008 (UTC)

Exponential applification

A see some changes based on the question of whether PCR produces exponential amplification. I believe it does, by most methods. An exception is the QuikChange method, where amplification is linear, as the extended primers don't overlap. QuikChange is limited by either the number of templates, or the number of primers.

Question. Does the QuikChange method correspond to one of the "Variations on the basic PCR technique"? Agesworth (talk) 08:14, 2 April 2008 (UTC)

Can you provide a link to a description of QuikChange PCR? My initial googling indicates this is a mutagenesis kit; while it uses PCR to generate the target fragment I can't see any indications that this is different from a normal PCR. Maybe it's tagged PCR? (That doesn't seem to be on the list; could be added.) But this would still be exponential. Madeleine ✉ ✍ 12:47, 2 April 2008 (UTC)

Yes, QuikChange is a commercial mutagenesis kit, but the concept and method is so simple that you certainly don’t buy “the kit” more than once. It is also known as “whole plasmid PCR” (The earliest reference I have is: Anne Hemsley, Norman Arnheim, Michael Dennis Toney, Gino Cortopassi, and David J. Galas A simple method for site-directed mutagenesis using the polymerase chain reaction Nucl. Acids Res. (1989) 17: 6545-6551; doi:10.1093/nar/17.16.6545). Originally they used Taq, but now we use Pfu.

Your two primers are substantially complementary. If primer pairs anneal, there is pretty much nothing to extend. Where a primer anneals to the plasmid, it is extended all the way around until the polymerase reaches the 5’ end of the primer, where polymerisation stops. It has replicated one strand of the plasmid, with a remaining nick, and perhaps a missing base or two at the nick. The reverse primer does the same thing, except that the nick is in a different place, 30-40 basepairs away. When the two strands anneal together, you have a doubly nicked plasmid, but the overhang is so large that it is very stable, transforms well, and the two nicks are repaired by the cell’s repair DNA repair mechanisms. Thus, the method is also an example ligation free subcloning.

As the primer extensions don’t contain sequence complementary to the other primer, primers can’t anneal to primer extensions. Polymerisation can only occur when a primer anneals to the plasmid template. The reaction is therefore not exponential/geometric, but linear. Compared to normal PCR, the amount of template used is large, and the number of cycles is small.

All of this is detailed in the product description, eg http://www.chemistry.montana.edu/~martint/BCHM444/QuickChange.Mutagenesis.pdf (a google result).

What’s not clear to me is whether QuikChange is really “PCR”. It uses a PCR machine, and all of the standard reagents and methods, but as I said, primers don’t anneal to primer extensions, and so the reaction is not actually a “chain” reaction. I don’t know what “tagged PCR” is. Agesworth 23:51, 2 April 2008 (UTC)

Tagged PCR is different, it's simply adding additional sequence to the 5' end of primers; these sequences can be used for a later amplification from the added sequence, or whatever else you want...

I think you're right, I wouldn't describe this as PCR... it's not even intended as DNA amplification, really. Other DNA amplification strategies that aren't PCR are rolling circle (also linear) and exponential/hyperbranched rolling circle amplification. (Interestingly, these all involve circular templates.) I can't think what article this would be appropriate to, but it's so specific it doesn't seem important to cover it.Madeleine ✉ ✍ 00:05, 3 April 2008 (UTC)

Lead picture change

The new pic is hard to read and is copyrighted-- and this is is not fair use. I've got an obvious bias for thinking so, but I thought the old pic was fine. I was pleased to see it used elsewhere: [5] -- Madeleine ✉ ✍ 00:41, 8 April 2008 (UTC)

I don't know your bias, but I agree, the picture was not suitable for multiple reasons. --Agesworth 02:07, 8 April 2008 (UTC) —Preceding unsigned comment added by Agesworth (talk • contribs)

It's my hand. :-) Madeleine ✉ ✍ 03:56, 8 April 2008 (UTC)

If the pic is indeed copyrighted it preempts any rationale for use here, and I agree that it's not very appealing to the eye. It would be nice though to have a pictorial summary diagram of the PCR process, especially alongside the lead. The "hand" picture could still be in there, though ;-). Malljaja (talk) 10:10, 8 April 2008 (UTC)

Well, I made that PCR diagram (figure 2) farther down in the page. If it could be improved at all just let me know. I think it's too big for the lead, though. I could try making a version of that that only depicts the double-stranded molecules after each step, which would make it a bit smaller. Point out a link if you see a pic you like online that would be good.

Would you worry about such a pic being redundant with the more detailed version fig 2? Madeleine ✉ ✍ 13:21, 8 April 2008 (UTC)

The new and now deleted pic encapsulated the basic principle of PCR well, in that it visually connected the DNA double helix with the PCR process, which is great for a layperson audience. Figure 2 is very good too, but it requires some knowledge to fully get one's head around it--eg there's no explanation as to what the individual bars and errors represent. A more detailed legend would be starting point, but a linkage with the structure of DNA that now everyone recognises would really top it off. So a fleshed-out version of figure 2 would be excellent for the lead. So that would be my 2 cs. I don't want to foist this on you though--if you have time (and drawing tools) to redo the figure that would be great, otherwise we do it perhaps at small increments. Thanks!Malljaja (talk) 14:37, 8 April 2008 (UTC)

Bars? Errors? The picture has elements in it not explained in the text that seem important but could be removed: the polymerase (green circle) is depicted in the first annealing and extension steps, the template DNA is blue, the primers are red, and newly synthesized DNA is green. Because this is an SVG, it's fairly easy to delete things or change the color of things. Which, if any, of these items should be removed?

It sounds like you'd like something at the top that depicts the unraveling of a DNA double helix to two lines, to explain that the lines are representing paired DNA strands. This could be added at the top, maybe.

I think a PCR diagram is necessarily information dense. Note that the copyrighted image that started this discussion is so dense that it would take up the entire width of the page to become readable. To avoid this level of density I had avoided trying to labeling everything in the diagram I made. It also inherits a style from the diagram I was copying/correcting [6]. Madeleine ✉ ✍ 15:56, 8 April 2008 (UTC)

My apologies, that should have been "lines and arrows"--I've been working too much on graphs these days... I'd expect a general audience might be a little flummoxed by the lines and arrows in much the same way as I am, when I'm looking at some arcane chemical drawings. So how about expanding the legend by including what the lines, circles, etc represent? I agree though that it's going to be a little bit of a challenge to accommodate all the other details I suggested. The DNA helix I envision would be rather small. I'm rather hamfisted when it come to drawing, so I'm not going to touch it myself for now. Malljaja (talk) 16:15, 8 April 2008 (UTC)

AfD Polymerase Chain Reaction (simplified)

There is currently a simplified version of this article, Polymerase Chain Reaction (simplified), which is being considered for deletion at Wikipedia:Articles for deletion/Polymerase Chain Reaction (simplified). - Eldereft ~(s)talk~ 22:25, 5 May 2008 (UTC)

Closed - deleted as unnecessary, at least for the nonce. - Eldereft ~(s)talk~ 07:29, 11 May 2008 (UTC)

Incorrect sentence

"Two primers, which are complementary to the DNA regions at the 5' (five prime) or 3' (three prime) ends of the DNA region."

This is false. Both primers are complementaty to 3' end, beacause they must have orientation 5'->3' to be primers. The main difference between them is the strand that they are complemantary to. One bind with sense, and the other with antisense strand. —Preceding unsigned comment added by 217.173.184.64 (talk) 14:09, 3 May 2009 (UTC)

Thanks for catching this--I've modified this section to reflect that fact. Malljaja (talk) 15:59, 3 May 2009 (UTC)

PCR principles and procedure: Melting

I changed "melting" to separation to more accurately define the process. DanielGlazer (talk) 21:47, 13 June 2009 (UTC)

Thanks, though "melting" is used in scientific literature to describe the separation of DNA strands due to heat. "Separation" is probably a better term to use in an article for a general audience, though. Adrian J. Hunter^{(talk•contribs)} 01:59, 14 June 2009 (UTC)

Crystal structure of hexagonal ice. Gray dashed lines indicate hydrogen bonds

3 HB binding GC

2 HB binding AT

Melting is specific while separation not. Why do you think that imaginary 'general audience' attention should be separated to nowhere? The kinetic energy of molecules in PCR reaction environment increase with temperature. The DNA hydrogen bonds dissociate. This is parallel (both verbally and conceptually) with melting of ice when hydrogen bond becoming to week to hold solid state structure, the point when water is melting. What do you think to put back melting and link to [DNA melting]? Xook1kai Choa6aur (talk) 17:52, 30 August 2009 (UTC)

This seems reasonable to me. – ClockworkSoul 00:07, 31 August 2009 (UTC)

I also prefer the term "melting", though I recognise that this term is potentially confusing for a lay or beginners' audience. The analogy with melting of water ice, while intuitive, may invite the misconception that DNA "dissolves" into very small molecules rather than separates into its two strands. The lede states "These thermal cycling steps are necessary to physically separate the strands (at high temperatures) in a DNA double helix (DNA melting)", and in the description of the PCR steps it says "It causes separation of DNA template and primers by disrupting the hydrogen bonds between complementary bases of the DNA strands, yielding single strands of DNA." I suggest to reword these two sentences to "These thermal cycling steps are necessary to physically separate the two strands in a DNA double helix at high temperatures in a process called DNA melting,..." and to "It causes separation of the DNA template by disrupting the hydrogen bonds between complementary bases, yielding single strands of DNA." The latter sentences also fixes a small error that suggested separation of the primers during denaturation. Malljaja (talk) 02:46, 31 August 2009 (UTC)

(M:fixes a small error that suggested separation of the primers during denaturation X:Primers melts before dsDNA melts. Which error?)

Not separation which in chemistry has distinctive meaning it is purification/mass transfer processes. If rely is need then dissociation may be better. See also Dissociation constant for water and DNA. Instead of using descriptive imprecise (what is somehow misleading) language isn't be better for introductory explanation to draw a picture?

By the way this article do not mention low temperature T4 polymerase added after each melting cycle in early days. The "chain reaction" is basically imprecise (Cetus advertising?) since the reaction has discrete controlled steps.

Here is the stepped math to add:

If N cycles were set then: the number of of new, amplified particles (Ap) to the number of particle in sample(Sp) is not greater than sample binary exponent.

• Sp ≤ Ap*2^N

Xook1kai Choa6aur (talk) 03:32, 31 August 2009 (UTC)

(I changed the indenting above.)

Actually my original concern with "melting" was that it suggested the DNA began as a solid, rather than in solution. But Malljaja's proposed wording makes things very clear, spelling things out in a way that avoids confusion with either solid/liquid transition or mass transfer.

Regarding "chain reaction", I don't follow XC's reasoning. To me chain reaction simply implies that the products of one reaction become the substrate for the next reaction. I'm sure this term is used in other descriptions of PCR that have no affiliation with Cetus. Adrian J. Hunter^{(talk•contribs)} 14:41, 31 August 2009 (UTC)

Without melting, after each extension step, the reaction will self terminate. There is no self chaining but externally controlled thermal ssDNA regeneration. It is slight distinction, to chained self propelled reaction. Distinction worth additional word - stepped. After searching for US patent 4,683,202[7] one can read inside :The steps of the reaction may be carried stepwise and can be repeated..., the process of reaction utilizes repetitive reaction(1-15:20) each of single strands produced in step (b)(3-5:10) the steps may be conducted...in addition step (b) and (c)(3-10:15) in general the process involves chain reaction for producing, in exponential quantities relative to the number of reaction step involved(5-20:25) . Are the word "stepped chain reaction", as added here "vague/misleading/unnecessary"?

Double standard DNA is rigid, it behaves as stiff rod. Melted do not behave as stiff rod. Molecular kinematic of DNA may change between solid-fluid like properties.

About the separation process of ssDNA how one may know that melted DNA single strands are separated. Booth ssDNA can form in volume space knob or any other string structure; being melted, but more space congested, closer to each other, than before melting; thus before that proposed 'separation'/melting (What is clearly illogical that after 'separation' the particles are closer). The word here is very unfortunate choice, dissociation is better. Its origin may point to DNA denaturation, where urea context, clearly mark mistaken literal source understanding; since denaturated ssDNA is separated in the mass transfer/purification context on chromatographic columns in urea buffer. Urea buffer also denturate/melt DNA. See links added in talk DNA denaturation. Xook1kai Choa6aur (talk) 06:04, 1 September 2009 (UTC)

Infos about its SOPs and compliance issues.....

• http://scholar.google.com.hk/scholar?hl=en&safe=strict&q=polymerase%20chain%20reaction%20standard%20operating%20procedures&um=1&ie=UTF-8&sa=N&tab=ws
• http://www.google.com.hk/search?hl=en&safe=strict&q=polymerase%20chain%20reaction%20standard%20operating%20procedures&um=1&ie=UTF-8&sa=N&tab=sw

• http://scholar.google.com.hk/scholar?hl=en&q=polymerase+chain+reaction+compliant&btnG=Search&as_sdt=2000&as_ylo=&as_vis=0
• http://www.google.com.hk/search?hl=en&q=polymerase%20chain%20reaction%20compliant&sa=N&tab=sw

• http://scholar.google.com.hk/scholar?hl=en&q=polymerase+chain+reaction+compliance&btnG=Search&as_sdt=2000&as_ylo=&as_vis=0
• http://www.google.com.hk/search?hl=en&q=polymerase%20chain%20reaction%20compliance&sa=N&tab=sw

--222.64.222.45 (talk) 04:54, 11 June 2010 (UTC)

Infos about its inactivation.....

• http://scholar.google.com.hk/scholar?as_q=polymerase+chain+reaction+inactivation&num=10&btnG=Search+Scholar&as_epq=&as_oq=&as_eq=&as_occt=title&as_sauthors=&as_publication=&as_ylo=&as_yhi=&as_sdt=1.&as_sdtp=on&as_sdts=5&hl=en

--222.64.222.45 (talk) 05:12, 11 June 2010 (UTC)

Nucleoside or nucleotide

A "nucleotide" is a 5'-phosphate ester of a nucleoside. A "nucleoside" results from the linkage of a ribose or deoxyribose sugar with a purine or pyrimidine base through an N-glycosidic linkage. The article refers to nucleotides throughout. The wiki articles on Nucleotides and Nucleosides also follow this convention; the PCR page does not follow suit. The "TP" in dNTP clearly refers to "triposphate", therefore we have a phosphorylated nucleoside (which is known as a "nucleotide"). The usage of "nucleoside" is not technically correct - even though the error is widely made. If this article insists on referring to dNTPs as nucleosides, then you'd better fix up all the dozen or so other articles that use that terminology including this one where nucleotides are mentioned several times.

The line: "Deoxynucleoside triphosphates (dNTPs; also very commonly and erroneously called deoxynucleotide triphosphates)" should read "Deoxynucleotide triphosphates (dNTPs; also very commonly and erroneously called deoxynucleoside triphosphates)"

—Preceding unsigned comment added by 88.104.225.28 (talk) 19:11, 21 January 2011 (UTC)

Thanks for clarifying your change. This perpetual confusion seems to arise because of the moniker, nucleotide triphosphate, which is a little problematic (the term nucleotide comprises the base, ribose, and phosphate, so the added triphosphate behind nucleotide is tautological). However, (deoxy)nucleotide triphosphate is in very common usage in molecular biology and differentiates the chemical from the mono- and diphosphate forms. So I've reverted the entry to your preferred description with which I now agree. If it's not too much trouble, I'd suggest that you create an account, because such small changes by IPs are more likely to be reverted. Also, please consider adding a short explanation for your changes into the Edit summary box—that way it indicates that the aim of your edit is to improve and not to do mischievous damage to an entry. Thanks. Malljaja (talk) 21:04, 21 January 2011 (UTC)

I agree with the first paragraph by 88.104.225.28 but not the second. I've always understood that a dNTP can either be referred to as a "nucleotide" or, more specifically, as a "nucleoside triphosphate"; I believe the term "nucleotide triphosphate" to be incorrect (though common). The term (deoxy)nucleotide triphosphate is not needed to distinguish from mono- or diphosphate terms: that's properly achieved by the "tri" in "nucleoside triphosphate". I've checked my organic chem/biochem textbooks at home but none are as clear as they could be on this point. Presumably the source that would clear this up is Nucleoside and Nucleotide Nomenclature, though I can't access it from home. Adrian J. Hunter^{(talk•contribs)} 11:31, 22 January 2011 (UTC)

I got the source linked above through the WikiProject Resource Exchange but it wasn't helpful, nor could I find anything useful trying to search IUPAC. But I did find a few other lines of evidence:

• Hits for various searches on NCBI:

search	"nucleoside triphosphate"	"nucleotide triphosphate"	excess
PubMed – abstract, keywords or title	1506	302	5-fold
PubMed – title only	356	30	12-fold
protein	334346	4594	72-fold

The trend is that the more scrutiny an instance of either term is likely to receive, the more likely it is to be "nucleoside triphosphate", rather than "nucleotide triphosphate".

• Of all the biology textbooks that are freely searchable through NCBI ([8]), several contain eight or fewer instances of either term, and many of these contain both terms. Only two books have more than eight instances: Biochemistry. 5th ed. has 22 instances of "nucleoside triphosphate" and 1 of "nucleotide triphosphate"; Molecular Biology of the Cell. 4th ed.—a very highly regarded textbook—uses exclusively "nucleoside triphosphate" (19 instances).
• Terms such as "adenosine triphosphate" arguably follow the "nucleoside triphosphate" pattern, as adenosine is the name of a nucleoside.

So I couldn't find any evidence that either term was incorrect, but there's plenty of evidence that "nucleoside triphosphate" is the preferred term. I've changed the article accordingly.

Adrian J. Hunter^{(talk•contribs)} 14:23, 23 April 2011 (UTC)

I had a hard time distinguishing "deoxynucleotide" in "sometimes called "deoxynucleotide triphosphates" from "deoxynucleoside" at the beginning of the entry. Is it feasible to italicize the "tide" in "deoxynucleotide" to emphasize the difference? V35b (talk) 23:11, 22 October 2023 (UTC)

Thanks, Adrian, for looking into this with such an exceedingly fine comb. Very good work. Though I do not need to be convinced that the "nucleoside" is technically the correct term, I foresee that some future editors may revert this back to "nucleotide" because of the vagaries of informal usage of this term. How about mentioning the alternative usage (e.g., as "also sometimes called deoxynucleotides")? I've re-linked the term, to remove the glaring red link, which may invite another revert back-and-forth. Many thanks! Malljaja (talk) 02:13, 24 April 2011 (UTC)

Too much free time over Easter, I guess . I've implemented your sensible suggestion in a manner that I hope clarifies the relationship and the wikilink target, along with a <!-- note pointing to this discussion. Adrian J. Hunter^{(talk•contribs)} 09:11, 24 April 2011 (UTC)

Selective DNA isolation

The article mentions this many times, but it doesn't seem particularly clear to me. Does it mean the DNA alone is amplified, or that specific parts of the genome can be chosen to be amplified specifically? If it's the latter (which it seems to be referring to), how? I've spent the last 20 minutes looking for an explanation, but unless I'm missing something obvious, there isn't one. 86.183.150.203 (talk) 12:32, 10 March 2012 (UTC)

Removal of circumstances of discovery - LSD

I would like to take note of the reverting of this page, removing my contribution to the history section, about the discovery of PCR through the usage of LSD. Malljaja reverted the article for the following reason: "This entry is about PCR not K Mullis.". My contribution was about the rest of the story behind the discovery, which is conspicuously left out, not about Mullis, per se: "Years after his work on the development of PCR, Mullis admitted in private and publically that his experimentation with LSD was a major influence on his work. During a symposium held for centenarian Albert Hofmann, "Hofmann revealed that he was told by Nobel-prize-winning chemist Kary Mullis that LSD had helped him develop the polymerase chain reaction that helps amplify specific DNA sequences."[51] When questioned about this during an interview for BBC's Psychedelic Science documentary, Mullis stated "What if I had not taken LSD ever; would I have still invented PCR?" He replied, "I don't know. I doubt it. I seriously doubt it."[52]"

If this is to be deleted, you may as well also delete this part, which also includes facts about Mullis' life (and PCR as well):

"When Mullis developed the PCR in 1983, he was working in Emeryville, California for Cetus Corporation, one of the first biotechnology companies. There, he was responsible for synthesizing short chains of DNA. Mullis has written that he conceived of PCR while cruising along the Pacific Coast Highway one night in his car.[49] He was playing in his mind with a new way of analyzing changes (mutations) in DNA when he realized that he had instead invented a method of amplifying any DNA region through repeated cycles of duplication driven by DNA polymerase."

Perhaps the formatting (essentially a copy and paste from the article on Mullis) is badly done, but I really believe the fact that PCR was discovered as a direct result of LSD should be included in the history section of the article. Any suggestions on how to make this happen? — Preceding unsigned comment added by Tr0798 (talk • contribs) 02:16, 12 May 2013 (UTC)

You're not the first trying to add this info to this entry. Time permitting I can try to locate the relevant discussion that ensued at that time. Suffice it to say for now, there's not much information in this bit you added—K Mullis is a very colourful character, and having this info in his entry is fine. However, most scientists who are familiar with the history of PCR are very aware that the technique was neither "invented" by Mullis nor the result of LSD-induced hallucinations. As it says in the entry, other researchers (notably Kleppe and colleagues) had laid the theoretical groundwork, and the advent of heat-stable polymerases set in motion technological advances that led to PCR—Mullis' only achievement in this was to milk a great story out of it, one that has gained some notoriety but not found wide acceptance in the field of molecular biology. Malljaja (talk) 02:53, 13 May 2013 (UTC)

Proposed merge with Applications of PCR

The applications of PCR article is short enough to be merged into the main PCR article, and would add relevant info to the main article. TYelliot | Talk | Contribs 15:16, 10 June 2013 (UTC)

The PCR article already mentions some applications and the separate applications article should be merged with these sections. Smischke (talk) 15:37, 1 August 2013 (UTC)

There are a lot of different applications for PCR and I feel combining the two articles would clutter the wiki page. Yes, the main PCR wiki page mentions some applications, as it should, but the reference to the Applications of PCR wiki page serves to continue the discussion and go into greater detail. --Triesault (talk) 18:07, 9 October 2013 (UTC)

The applications page seems too specific to warrant its own page. The article is only slightly longer than the section on the PCR page itself. It should be merged, and duplicate content should be fixed. Alexwho314 (talk) 05:18, 21 February 2014 (UTC)

Yes merging it is useful. Please merge to make it more coherent and avoid duplication. (Drsoumyadeepb (talk) 06:04, 26 May 2014 (UTC))

I prefer to take the long view and upgrade the other article and allow the knowledge base to grow out Wikidgood (talk) 00:38, 19 October 2014 (UTC)

The issue is too figure out just how much the applications page can be expanded. Currently, it would fit well in the main PCR article, but elaboration and more uses would make having a separate article worthwhile. Alexwho314 (talk) 03:51, 29 October 2014 (UTC)

The amount of elaboration and more uses won't add too much clutter to the main page. Right now the uses section of the main page is too short while the PCR uses page is not very long. — Preceding unsigned comment added by 2602:306:348C:A6B0:4D72:1FEA:4086:D5EE (talk) 23:08, 25 June 2015 (UTC)  Done

Photograph Captions in Error or Misleading

The captions of the first 2 photos have a common problem, stating (typically) :

"A strip of eight PCR tubes, each containing a 100 μl reaction mixture"

I think this is incorrect or misleading (depending on interpretation) because:

(1) The scale of the human hands clearly indicates a single tube unit would be far less than 100ml in volume) (2) Typically a single tube such as shown is 0.2, 0.5 or 1.0ml

This raises the question, was the caption intended to describe the volume of a single tube in the 8 tube set or the 8 tube set volume in aggregate?

In any case, the total would be far less than 100ml so this should be corrected and I hope the original content editor could go back to the source to verify the correct volumes or replace the photos with equivalents of known volume.

Thank you.    Xiao-zi : 小紫 15:25, 22 June 2013 (UTC)  — Preceding unsigned comment added by Xiao-zi (talk • contribs)  

Hi Xiao-zi, the captions actually refer to a microlitre (μl), which is one thousandth of a millilitre (mL). 100 μl is 0.1 mL, and looks consistent with the photographs. Adrian J. Hunter^{(talk•contribs)} 02:45, 24 June 2013 (UTC)

PCR application in electrical engineering

An electrical engineering team was looking at creating slightly bent artificial DNA. PCR is used to make many copies, which are covered in a thin gold layer. These are useful as circular polarizers in optical fibers. — Preceding unsigned comment added by 75.149.135.25 (talk) 18:57, 8 April 2014 (UTC)

Picture under "Procedure"

I'm talking about the picture where the denaturation, annealing and extension phases are shown. I know there used to be another picture here, like the one in the spanish pages, that did not look as fancy. The current picture does not show any PCR-products because these are not made until the third cycle. I suggest using the older picture that is also in the spanish section, or make a new one that actually shows the first 3 cycles of a PCR and thus some PCR-products.

Solveig — Preceding unsigned comment added by 2001:700:303:3:124:2B2B:B648:4B3C (talk) 09:43, 3 October 2014 (UTC)

The current picture is confusing with respect to length of the various fragments:
1.The initial double-stranded template has been cut off square outside of the primers. Is this necessary, or the template can be indefinite length as long as it includes both primer regions?
2. In the elongations step, the new strands are elongated beyond the ends of the template strands. Is this possible?
In principle the diagram showing 2 or more cycles like this should make it obvious why you get a defined-length product starting with template of arbitrary length. I realize this is schematic, but it is misleading. See also topic three down from this: "Erroneus . . ."
Eaberry (talk) 03:17, 18 April 2020 (UTC)

I created an updated version of the figure replacing the erroneus one:

• 
  New figure

1. A set of primers (forward and reverse primers) is designed to specifically bind to complementary DNA fragments. The sites bound by the primers (light blue in new figure) flank the region of interest (darker blue). The regions bound by the primers and everything in between is copied by PCR. In the new figure, grey squares represent the nucleotides (DNA parts) that are not of interest. Usually, samples used for PCR contain mostly these "grey" regions and the region of interest is much shorter in comparison (but for the figure the focus is on the region of interest, of course).

2. No, I fixed that.

3. 3rd and nth cylce were added in the new figure.

 Done Enzoklop (talk) 13:11, 12 November 2020 (UTC)

Vagueness Tag on Procedure Section

I'm an electrical engineer with very little knowledge of Biology. I read through the Procedure section, and it made sense to me; I don't think the Vagueness tag is warranted. Smithderek2000 (talk) 22:35, 18 November 2015 (UTC)

Agarose Percentage

It may be helpful to enhance this page by mentioning what different % Agarose can provide to the clarity for different sizes of genes that are run through a gel electrophoresis. — Preceding unsigned comment added by Arklipp (talk • contribs) 19:43, 10 February 2016 (UTC)

Thanks for the suggestion, but as PCR is a very big topic, I think such information is better contained at Gel electrophoresis and Agarose gel electrophoresis. Note PCR is not always followed by electrophoresis, nor is it always used for amplifying genes. Adrian J. Hunter^{(talk•contribs)} 01:24, 11 February 2016 (UTC)

Please replace erroneous scheme

I made a new one (see German Wikipedia). The present one shows elongation of the PCR beyond the template, which does not occur. I was technically not able to replace the scheme myself. — Preceding unsigned comment added by WiWiki (talk • contribs) 19:47, 12 November 2017 (UTC)

The German version seems much better- https://upload.wikimedia.org/wikipedia/commons/thumb/2/2d/PCR-Schema-v2.svg/842px-PCR-Schema-v2.svg.png . Eaberry (talk) 03:43, 18 April 2020 (UTC)

I created a new figure and corrected the issues. See above (Talk:Polymerase_chain_reaction#Picture_under_"Procedure").  Done Enzoklop (talk) 13:16, 12 November 2020 (UTC)

Repetitive Extragenic Palindromic Sequence-Based PCR Analysis

Should REP-PCR have a discussion? LeadSongDog come howl! 21:34, 11 June 2018 (UTC)

"Ct value"

Hi there,

apparently there exists a term "Ct value" relative to PCR testing (as in www.cdc.gov/coronavirus/2019-ncov/hcp/duration-isolation.html), but i can't find anything about that in here (the term is used, but unexplained, in the COVID-19 testing article). This NY Times article has me understand that "Ct" stands for "cycle threshold", apparently "the number of amplification cycles needed to find the virus". Shouldn't this term be present and explained in here ? —Jerome Potts (talk) 01:45, 6 September 2020 (UTC)

The method used for COVID-testing is variation of the standard PCR method: Real-time_polymerase_chain_reaction. The term Ct value is explained in that article (see Real-time_polymerase_chain_reaction#Modeling.  Done Enzoklop (talk) 10:18, 18 November 2020 (UTC)

A Commons file used on this page or its Wikidata item has been nominated for speedy deletion

The following Wikimedia Commons file used on this page or its Wikidata item has been nominated for speedy deletion:

• Assist-plus-transferring-mastermix.jpg

You can see the reason for deletion at the file description page linked above. —Community Tech bot (talk) 16:09, 28 July 2021 (UTC)

No mention of Covid 19

Why is there no mention of the use of PCR in Covid 10 virus — Preceding unsigned comment added by 74.221.71.11 (talk) 16:41, 4 August 2021 (UTC)

Perhaps because there is nothing special or unique about the PCR use in amplifying the SARS-CoV-2 sequence. — kashmīrī ^TALK 22:27, 4 August 2021 (UTC)

Semi-protected edit request on 29 August 2021 -

PCR Test 'diagnostic' is a misleading statement that make Wikipedia and authors liable for FRAUD and possibly contributing to higher crimes:

https://en.wikipedia.org/wiki/Kary_Mullis = doesn't mention CovID-19 testing...

https://en.wikipedia.org/wiki/COVID-19_testing#Reverse_transcription_polymerase_chain_reaction_test = should attempt to be impartial and balance 'claims' with other REPORTS and information such as https://walkinverse.com/dr-kary-mullis-slams-fauci/ https://www.zerohedge.com/medical/why-covid-19-testing-tragic-waste

https://en.wikipedia.org/wiki/Polymerase_chain_reaction = list PCR Test as a detection of pathogens in nucleic acid tests for the diagnosis of infectious diseases. Samsonia8800 (talk) 15:34, 29 August 2021 (UTC)

 Not done: it's not clear what changes you want to be made. Please mention the specific changes in a "change X to Y" format and provide a reliable source if appropriate. ScottishFinnishRadish (talk) 15:44, 29 August 2021 (UTC)

Infectious disease source

The source cited for the sentence about using PCR for infectious diseases caused by bacteria and viruses is an article that seems to only discuss bacterial infections in veterinary settings. Should the sentence be reworded to remove “viruses” or is there another source that can be cited? 64.224.250.45 (talk) 04:17, 23 December 2021 (UTC)

Request to edit Patent expiry information with expiration confirmation

The section on Patent Disputes is very misleading. Patent expiry information from an authoritative source is available. — Preceding unsigned comment added by Donnelt (talk • contribs) 16:29, 9 January 2022 (UTC)

Lights on but nobody home? Locking an entry is one thing but such autocratic decisions require greater attention to flexibility and challenge. Tom Donnelly (talk) 08:48, 13 January 2022 (UTC)

Pcr

Is it for Ag or AB 2001:16A2:FE2D:2800:132:CA84:863B:7A02 (talk) 22:12, 25 January 2022 (UTC)

Error in the "Tucker PCR" image used "Procedure" section.

At step 5 of the image, the direction of the red primer is incorrect. It should be 3' 5', not 5' 3'.

121.162.195.157 (talk) 05:49, 1 April 2022 (UTC)

I second that! Thank you - I am right now developing a very simple software model of this process and in straining to understand the whole sense/anti-sense thing I was totally thrown by that labeling. 140.32.108.251 (talk) 16:55, 26 July 2022 (UTC)

Semi-protected edit request on 5 April 2022

In the first sentence, please remove

billions of copies (complete copies or partial copies) of a specific

and add

billions of copies (complete or partial) of a specific

There's no need for three "copies" in one sentence, since the meaning is unambiguous. 49.198.51.54 (talk) 07:45, 5 April 2022 (UTC)

 Done Happy Editing--IAmChaos 12:10, 5 April 2022 (UTC)