An Overview of Cloning DNA: The Biological Revolution

Cloning has a lot of new terms and jargon. Here is a list of terms with which you should become somewhat familiar as you work through the following lessons.

cloning and subcloning, cDNA, oligo-dT purification of poly-A mRNA
vectors-- plasmids, cosmids, shuttle, expression, insert
ligation-- anneal sticky ends, blunt-end, directional cloning, in-frame with promoter, linkers, polylinker sequence
transformation-- competent cells, antibiotic selection, blue/white selection
DNA libraries-- genomic, cDNA, colony/plaque hydridization
Hybridizations-- Southern (DNA), northern (RNA), western (Protein), colony/plaque
Enzymes-- DNA polymerase, DNA ligase (T4), reverse transcriptase, Taq polymerase
Promoters-- inducers, fusion proteins, IPTG, X-gal, reporter gene, green fluorescent protein
PCR
In vitro mutagenesis
GMO/transgenic organism

Necessary Components for Cloning DNA [local] (greatly simplified):

• Restriction endonucleases
• Plasmids or other cloning vectors
• Bacterial transformation
• Selection of transformed bacteria containing plasmid
• Detection of desired recombinant DNA by hybridization or protein expression

Also necessary for cloning is DNA Ligase [local]. This enzyme will take two pieces of DNA which are held together by complementary sticky ends and join them together with a phosphodiester bond. Once a fragment of foreign DNA is spliced into a replicating vector, it is termed recombinant DNA (rDNA).