A genomic library is a complete collection of cloned DNA fragments that constitute the entire genome of an organism. It plays an important role in research. The two types of DNA libraries are – genomic DNA library and cDNA library. Here, let’s learn about the construction of a genomic DNA library in detail.
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What is a Genome?
A genome is an organism’s complete set of DNA, including all of its genes. Each genome contains all of the information needed to build and maintain that organism. In humans, a copy of the entire genome is more than 3 billion DNA base pairs which are present in all cell nuclei. The nuclear genome contains protein-coding and non-coding genes, as well as other junk DNA and functional regions of the genome.
Most eukaryotes are diploid, which means that each chromosome has two copies in the nucleus, but the term genome refers to just one copy of each chromosome.
Genomic DNA Library
A genomic library or gene bank is a complete collection of cloned DNA fragments that constitutes the entire genome of an organism. It represents all the genes – expressed, non-expressed, intron, exons, etc. Genomic libraries can be kept for many years and the copies can be used for research purposes.
Advantages of a Genomic Library
- Genomic libraries derived from eukaryotic organisms are critical for studying the genome sequence of a particular gene of interest.
- It is useful for prokaryotes with small genomes to identify a clone encoding a specific gene of interest.
- It helps researchers to explore more about an organism’s genomic structure and function. It is also used to study genetic mutations.
- Pharmaceutically important genes can also be identified by this method.
Disadvantages of a Genomic Library
- It requires sophisticated software and a vast amount of computing power. Also, the process is prone to errors.
- Eukaryotic genome libraries with very large genomes contain many DNA that do not code for proteins, as well as non-coding DNA like repetitive DNA and regulatory regions, making them less than ideal.
Also see: Difference between cDNA and Genomic DNA Library
Construction of a Genomic DNA Library
The Genomic Library or gene bank is constructed by a shotgun experiment where the entire genome of the cell is cloned in the form of random and unidentifiable clones. It uses the chain termination method (Sanger’s sequencing) to sequence the DNA molecules. The DNA clones are produced by the following steps:
- Isolating the DNA fragments that are to be cloned and joining them with suitable vectors like the lambda phage.
- Now, it is introduced into the host cell at high efficiency to get a large number of independent clones.
- Finally, the desired clones are selected and used for the construction of the genomic library.
Steps to Construct a Genomic DNA Library
- First, the purification of the desired eukaryotic cell nuclei is done and this is accomplished through protease digestion and organic extraction.
- The derived genomic DNA is too large to be incorporated into a vector and must be fragmented into desired sizes. Both physical and enzymatic methods can be used to fragment DNA.
- There are several vectors available for cloning large DNA fragments. Phage, bacterial artificial chromosome, yeast artificial chromosome, and other such vectors are suitable for larger DNA. The λ replacement vectors are the most preferred ones for the construction of a genomic DNA library.
- Usually, the T4 DNA ligase enzyme is used to ligate the chosen DNA sequence into the vector.
- The recombinant vectors and the insert combinations are grown in a bacterial host cell (E. coli). They replicate their genome along with the vector genome contained within them.
- The collection of clones that contain all the sequences from the original source (including the sequence of interest) forms the gene bank or genomic library.
Screening of Clone
Each transformed bacterial host cell in a library will have only one vector with one DNA insert. The entire library can be plated over media on a filter. The filter, as well as colonies, are then hybridised, labelled with a probe and identified using detection methods such as autoradiography. Other techniques like PCR and the blue-white selection method can also be used to screen the clone.
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