Animal Biotechnology

PHAGES

• Phages are derivatives of bacteriophage lambda (λ phage), a virus which infects E. coli.
• They are linear DNA molecules, whose region can be replaced with foreign DNA without disrupting its life cycle.
• The major advantage of the λ phage vector is its high transformation efficiency, which is about 1000 times greater than that of the plasmid vector. They also have a larger cloning limit than plasmids, consisting of 8-25 kb.

Phage vector

• Lambda phage vector
  • Lambda phages are viruses that can infect bacteria.  The major advantage of the phage vector is its high transformation efficiency, about 1000 times more efficient than the plasmid vector.
  • The extreme ends of the lambda DNA are known as COS sites, each is single stranded, 12 nucleotides long. 
  • Because their sequences are complementary to each other, one end of lambda DNA may base-pair with the other end of a different lambda DNA, forming concatemers. 
  • The two ends of a lambda DNA may also bind together, forming a circular DNA. 
  • In the host cell, the lambda DNA circularizes because ligase may seal the join of the COS sites.
  • In the assembly process of lambda virions, two proteins Nu1 and A can recognize the COS site, directing the insertion of the lambda DNA between them into an empty head. 
  • The filled head is then attached to the tail, forming a complete lambda virion. 
  • The whole process normally takes place in the host cell.  However, to prepare the lambda virion carrying recombinant lambda DNA, the following in vitro assembly system is commonly used.
  • Proteins Nu1 and A are encoded by the genes in the lambda genome. If the two genes are mutated, lambda DNA cannot be packaged into the pre-assembled head.  Because tails attach only to filled heads, the cell will accumulate separate empty heads and tails, which can then be extracted.  When the extract is mixed with recombinant lambda DNA and proteins Nu1 and A, the complete lambda virion carrying recombinant lambda
  • DNA will be assembled.