1-2.2.1 Choice of host organism:

A good host should have the following properties:

• Easy to grow and transform.
• Do not hinder replication of recombinant vector.
• Do not have restriction and methylase activities.
• Deficient in recombination function so that the introduced recombinant vector is not altered.
• Easily retrievable from the transformed host.

Various hosts are used in rDNA technology depending on the goal: For example bacteria, yeast, plant cells, animal cells, whole plants and animals.

•  Prokaryotic systems such as E. coli are commonly used due to various advantages like, They have-

• Well studied expression system,
• Compact genome,
• Versatile,
• Easy to transform,
• Widely available, and
• Rapid growth of recombinant organisms with minimal equipment.

Only disadvantage is that they lack post-translational modification (PTMs) machinery required for eukaryotic proteins.

•  Eukaryotic systems are difficult to handle in contrast to bacterial hosts. They are favoured for expression of recombinant proteins which require post translational modification and only if they can grow easily in continuous culture.

1-2.2.2 Choice of vector:

Vector is an autonomously replicating (inside a host cell) DNA molecule designed from a plasmid or phage DNA to carry a foreign DNA inside the host cell. Transformation vectors are of two types:

• Cloning vector is used increasing the number of copies of a cloned DNA fragment.
• Expression vector is used for expression of foreign gene into a protein.
• If a vector is designed to perform equally in two different hosts, it is called a shuttle vector.

Properties of an ideal vector: A good vector should have the following characteristics:

• Autonomously replicating i.e. should have ori (origin of replication) region.
• Contain at least one selectable marker e. g. gene for antibiotic resistance.
• May contain a scorable marker (β -galactosidase, green fluorescent protein etc.)
• Presence of unique restriction enzyme site.
• Have multiple cloning sites.
• Preferably small in size and easy to handle.
• Relaxed control of replication to obtain multiple copies.
• Presence of appropriate regulatory elements for expression of foreign gene.
• High copy number

The selection of a suitable vector system depends mainly on the size limit of insert DNA and the type of host intended for cloning or expression of foreign DNA.