Reverse Transcriptase and Gene Therapy in Molecular Biology

Reverse Transcriptase

Production of DNA from RNA

Reverse transcriptase is an enzyme isolated from retroviruses that catalyzes the production of DNA from an RNA template.

Use in Molecular Biology

• Reverse transcriptase is used in recombinant DNA technology to produce genes for gene transfer.
• It catalyzes the production of complementary DNA (cDNA) from an mRNA template.
• cDNA does not contain introns because mRNA undergoes splicing before cDNA production.
• Bacteria lack the machinery for intron removal, so genes inserted into them need introns removed to produce functional proteins.
• Examples of reverse transcriptase use include:
  • Mass production of human insulin by E. coli.
  • Generation of cDNA libraries for DNA microarrays (DNA fingerprinting).

Gene Therapy

Somatic vs. Germ Line Therapy

Gene therapy inserts genes into an individual's cells to treat genetic diseases.

Somatic Therapy: Genes inserted into body cells; modification restricted to the individual patient; considered safe.

Germ Line Therapy: Genes inserted into germ cells; modification inherited by offspring; not considered safe.

Use of Viral Vectors

• Viral vectors replace defective genes with functional copies.
• Individuals with severe combined immunodeficiency (SCID) may be unable to synthesize adenosine deaminase (ADA).
• White blood cells or bone marrow cells are removed, and a normal gene copy is integrated using a viral vector.
• When the cells are replaced, the normal gene is expressed, producing ADA and treating SCID.
• Technical challenges remain, such as ensuring the correct amount at the right time and place.