In this article we will discuss about the genomic and cDNA libraries.
Genomic DNA Library:
A genomic library is a collection of independently isolated vector linked DNA fragments derived from a single organism. It contains at least one copy of every DNA sequence in the genome. An ideal library is one that represents all of the sequences with smallest possible number of clones.
The genomic DNA libraries can be prepared by the complete digestion of the total genomic DNA with a restriction enzyme and the fragments are inserted into a suitable vector like X phages (Fig. 22.11). The drawback of this method is that sometimes the sequence of interest may contain multiple restriction sites, so digestion with RE results into two or more pieces.
In this method the eukaryotic DNA is broken up into smaller fragments, thus an entire library would necessarily contain a large number of phages, and screening of which is very laborious. The problems of this method can be avoided by random shearing of total DNA and cloning of large fragments.
This method ensures that sequences are not excluded from the cloned library simply because of the distribution of restriction sites. In this procedure the randomly fragmented DNA is partially digested with RE which has short recognition sites. The fragments of desired size are collected through agarose gel electrophoresis, so the population of overlapping fragments that are close to random can be cloned directly.
Complementary DNA (cDNA) libraries can also be prepared by isolating mRNAs from tissues which are actively synthesizing proteins, like roots and leaves in plants, ovaries or reticulocytes in mammals, etc. The mRNAs are used for copying them into cDNAs through the use of reverse transcriptase. Then the cDNA molecule can be made double stranded and cloned (Fig. 22.12).
However, cDNA clones will differ from genomic clones in lacking the introns present in split genes, and have the advantage of being capable to be expressed in bacteria, which do not have the machinery to process the eukaryotic mRNA. There are far less number of cDNA clones in a bank than in a genomic library, which makes easier to look for a desired gene. Screening of cDNA bank also provides fairly unambiguous results.