Vectors
The term vector refers to the DNA molecules that act as transporting vehicle which carries target DNA into a host cell for the purpose of cloning and expression. Cloning vectors are used to clone target DNA whereas expression vectors are engineered so that desirable target DNA can be transcribed in RNA and translated into protein, A viral DNA or plasmid is generally used as a vector. The important features of a cloning vector are as follows:
1. Ability to replicate in host cells. All cloning vectors have origin of replication for autonomous replication within the host cell. The origin of replication is a specific sequence in DNA from where replication starts. When target DNA is linked to vector containing origin of replication then along with vector replication, desirable target DNA also starts replicating within the host cell.
2. Unique restriction sites for insertional cloning, All cloning vectors have features that allow a target DNA to be conveniently inserted into the vector. This may be a multiple cloning site (also called polylinker site), which contains many unique restriction sites. The restriction site(s) in the polylinker site are first cleaved by specific restriction enzyme(s), and a target gene is then ligated into the vector using DNA ligase.
3. Genetic marker to select for host cells containing the vector Genetic marker is a gene that allow the selection of transformed cell from non trans formed cells and recombinant containing transformed cells from non-recombinant containing transformed cells. Marker genes belong to two broad categories: selectable markers and screen able markers. A selectable marker gene encodes a product that allows the growth of one type of cells under specific conditions that kill or restrict the growth of other types of cells. A screenable marker gene, also called reporter gene, gives a product that can be detected using a simple and often quantitative assay.
4. Low molecular weight (minimum amount of non-essential DNA). The advantages of a low molecular weight are several. First, the plasmid is more resistant to damage by shearing and is readily isolated from host cells. Secondly, low molecular weight plasmids are usually present as multiple copies. Finally, with a low molecular weight there is less chance that the vector will have multiple sites for any restriction endonuclease.
