Principal methods used in molecular genetics

Principal methods used in molecular genetics

Brain mRNAs
Reverse transcriptase transcribes these different mRNAs into single complementary strands of DNA

Single-stranded brain cDNAs formed
Single-stranded cDNAs form template for double-stranded DNA

Double-stranded brain cDNAs formed
Double-stranded cDNAs added to bacterial plasmids that insert the cDNA into the plasmid

Recombinant DNA plasmid containing the brain DNA
Recombinant DNA then inserted into bacteria which reproduce the brain DNA

cDNA library formed with each bacterium multiplying the specific cDNA it contains

Individual bacteria isolated and cultured to produce clones that yield
specific cDNAs
Specific cDNAs (for example a receptor) inserted in plasmid that transfects a mammalian cell (e.g. fibroblasts) in culture

Mammalian cell containing the specific cDNA then exposed in a culture medium to a toxin which destroys all non-transformed mammalian cells
Add radioligand that identifies the receptor of the mammalian cell

Identification and isolation of transformed cells which can then be cultured to provide unlimited quantities of the receptor protein