when amplification of gene interest using pcr can directly  produce large dna fragraments of our interest ..then y should it be ligated with a vector and cloned by inserting it into a host ??.....pls explain   if am anywhere wrong:

Asked by Rzainabafra - | 17th Nov, 2016, 08:16: AM

Expert Answer:

The technique of molecular cloning allowed the study of individual genes of living organisms. However, this technique was dependent on obtaining a relatively large quantity of pure DNA. This relied on the replication of the DNA of plasmids or other vectors during cell division of microorganisms.

In order to study individual genes or specific DNA regions of interest, it is often necessary to obtain a large quantity of nucleic acid for study. Researchers found it extremely laborious and difficult to obtain a specific DNA in quantity from the mass of genes present in a biological sample.

Rather than isolate a single copy of the target DNA from a large number of cells, it is often more useful to generate multiple copies of a target from a single molecule of DNA or mRNA, via an in vitro amplification method. A technique that amplifies DNA through a simple enzymatic reaction, called PCR or Polymerase Chain reaction was developed by Kary Mullis at that time which enabled scientists to make millions or even billions of copies of a DNA molecule in a very short time.

A large quantity of DNA obtained through PCR can in fact be used for detection of pathogens like HIV, diagnosis of specific mutations, DNA fingerprinting, diagnosis of plant pathogens, etc.

However, PCR has certain limitations. DNA polymerase is prone to error, which in turn causes mutations in the PCR fragments that are made. Additionally, the specificity of the PCR fragments can mutate to the template DNA, due to nonspecific binding of primers. Furthermore, prior information on the sequence is necessary in order to generate the primers. Hence, except for the need of large quantity of DNA, the rDNA technology of ligation, cloning and amplification is preferred over PCR.

Answered by Sheetal Kolte | 17th Nov, 2016, 09:55: AM

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