Biotechnology: Principles and Processes - Tools of Recombinant DNA Technology

Restriction Endonucleases: Known as 'molecular scissors', these enzymes cut DNA at specific palindromic sequences. $EcoRI$ is a primary example, recognizing the sequence $5'-GAATTC-3'$.

Naming Convention: The first letter comes from the genus ($E$ for $Escherichia$), the next two from the species ($co$ for $coli$), the fourth from the strain ($R$ for $RY13$), and Roman numerals indicate the order of discovery.

Gel Electrophoresis: DNA fragments are negatively charged due to the phosphate group ($PO_4^{3-}$). They migrate towards the anode ($+$) through an $Agarose$ gel matrix. Smaller fragments move faster and further.

Visualization: DNA fragments are stained with $Ethidium\ Bromide$ ($EtBr$) and exposed to $UV$ radiation to appear as bright orange bands.

Cloning Vectors: DNA molecules used as vehicles to carry foreign DNA into a host cell. Key features include an $ori$ (Origin of Replication), selectable markers like $amp^R$ or $tet^R$, and cloning sites.

Selectable Markers: These help in identifying and eliminating non-transformants. Common markers include genes conferring resistance to antibiotics like $Ampicillin$, $Chloramphenicol$, $Tetracycline$, or $Kanamycin$.

Insertional Inactivation: A method to identify recombinants by inactivating a gene (e.g., $lacZ$ gene) through the insertion of foreign DNA. Recombinants appear as white colonies, while non-recombinants appear blue in the presence of $X-gal$.

Competent Hosts: Cells treated with specific concentrations of divalent cations like $Ca^{2+}$ to increase the efficiency of DNA uptake. Methods include Heat Shock ($42^{\circ}C$), Micro-injection, and Biolistics (Gene Gun) using $Gold$ or $Tungsten$ particles.