The motive of both cloning (not expression cloning) and PCR is to amplify the gene of ineterst. But still, why cloning is preferred over PCR and even after amplifying your gene of interest through PCR, why you're supposed to go for ligation with vector and then transformation?
It very much depends on what experiments or analysis you intend to do, there is no simple answer to your question. PCR amplification alone will get you DNA related to the gene of interest, but you can't really do anything with it. It just generates some DNA. If all you want to do is sequence the DNA, then it is sufficient. However if you wish to propagate it, or manipulate it in any way, then you need to have it cloned into a plasmid. Oftentimes you would do both, amplify it to generate the DNA in order to clone it. But it all depends upon what experiment you intend to perform.
It very much depends on what experiments or analysis you intend to do, there is no simple answer to your question. PCR amplification alone will get you DNA related to the gene of interest, but you can't really do anything with it. It just generates some DNA. If all you want to do is sequence the DNA, then it is sufficient. However if you wish to propagate it, or manipulate it in any way, then you need to have it cloned into a plasmid. Oftentimes you would do both, amplify it to generate the DNA in order to clone it. But it all depends upon what experiment you intend to perform.
Michael gives a very good answer to your question. Additionally, even when using PCR prior to sequencing, you may be amplifying two alleles from a diploid organism which may differ in length (especially when amplifying intronic or intergenic regions). You cannot reliably interpret the sequences when this happens so cloning would be necessary to sequence one allelic copy.
- I cannot really add much to the above informative answers
- Except to say that if you wish to sequence a PCR product and require full length sequencing then cloning can provide an advantage for 2 reasons:
- Then of course you have advantages already mentioned souch as the fact that toy are dealing with one clone per colony and not potentailly a mixed PCR population and the fact that you can make as much of the inserts as you like by bacterial replication
- I will add that in the context of mixed populations and sequencing, this is an advanatge if you pick and prep multiple plasmids (plus inserts) if you wish to clearly wish to identify SNPs or your polulation might contain INDELS which would result in out of frame jumbled sequence from a mixed nascent PCR population but potentially a disadvantage for identifying a heterozygote from a quick sequencing screen; as in PCR sequence you would see the double peak but the rest of your sequence would be a monoclonal ladder
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