Gene and Protein Engineering-Recombinant DNA

Techniques central to recombinant DNA technology are:

1.cleavage of DNA at specific sites by restriction nucleases, greatly facilitating the isolation and manipulation of individual genes.

2. DNA ligation, making it possible to design and construct DNA molecules not found in nature

3.DNA cloning through the use of cloning vectors or PCR, generating many copies of identical molecules.

4.Nucleic acid hybridization allows specific sequences of DNA/RNA with great accuracy and sensitivity on the complementary base sequence.

5. Rapid determination of the sequence of nucleotides of any DNA, allowing identification of genes and their amino acid sequence.

6. Nucleic acid microarrays allow many hybridization reactions to take place simultaneously.

Restriction nucleases are purified by bacteria and cut DNA at specific sites defined by the local nucleotide sequence, cleaving a long double stranded DNA molecule into fragments of strictly defined sizes. Different restriction nucleases have different sequence specificities.

The specific sequence recognized is normally 4-8bp long. If these sequences are in the genome itself they are protected from cleavage by methylation at an A or C nucleotide. Some restriction…