4-4.1. Introduction

A genomic library is an organism specific collection of DNA covering the entire genome of an organism. It contains all DNA sequences such as expressed genes, non-expressed genes, exons and introns, promoter and terminator regions and intervening DNA sequences.

4-4.2. Construction of genomic library

Construction of a genomic DNA library involves isolation, purification and fragmentation of genomic DNA followedby cloning of the fragmented DNA using suitable vectors. The eukaryotic cell nuclei are purified by digestion with protease and organic (phenol-chloroform) extraction. The derived genomic DNA is too large to incorporate into a vector and needs to be broken up into desirable fragment sizes. Fragmentation of DNA can be achieved by physical method and enzymatic method. The library created contains representative copies of all DNA fragments present within the genome.

4-4.2.1. Mechanisms for cleaving DNA

(a)  Physical method

It involves mechanical shearing of genomic DNA using a narrow-gauge syringe needle or sonication to break up the DNA into suitable size fragments that can be cloned. Typically, an average DNA fragment size of about 20 kb is desirable for cloning into λ based vectors. DNA fragmentation is random which may result in variable sized DNA fragments. This method requires large quantities of DNA.