1 DNA technology 20 marker mamby

What are the roles of restriction endonucleases and methylases in bacterial DNA protection?

Restriction endonucleases cut DNA at specific sequences, while methylases protect bacterial DNA by methylating bases to prevent restriction enzyme activity.

How do restriction enzymes aid in cloning and genetic engineering?

Restriction enzymes create sticky or blunt ends, which are crucial for DNA ligation. Sticky ends facilitate joining complementary sequences, while blunt ends are less efficient for ligation without additional modifications.

What is the difference between genomic and cDNA libraries?

Genomic libraries contain DNA fragments representing an entire genome, while cDNA libraries contain only expressed genes, created by reverse transcribing mRNA into DNA.

What is the process for creating cDNA libraries?

Isolate mRNA, reverse transcribe it into DNA, and clone the cDNA into plasmids for bacterial transformation.

How is bacterial transformation performed, and why is it important for recombinant protein production?

Competent bacteria are treated with calcium chloride to make their cell walls permeable, allowing plasmid DNA uptake. This process enables gene expression and protein production.

How do viral vectors apply to vaccine development?

Viral vectors, such as adenoviruses, deliver genetic material (e.g., SARS-CoV-2 spike protein gene) to human cells to trigger an immune response, as seen in COVID-19 vaccines like Moderna.

What are recombinant proteins and how are they produced?

Recombinant proteins are produced by inserting a gene into a heterologous system (like bacteria or mammalian cells), which expresses proteins not naturally made by the host.

What features must plasmid vectors have to function effectively in gene cloning?

Plasmid vectors must include an origin of replication, a promoter for gene transcription, and a selectable marker for identifying successful clones.

What are the advantages of eukaryotic expression systems for protein production?

Eukaryotic systems are preferred for human proteins because they can perform post-translational modifications essential for proper protein function.

What role do bioreactors play in protein production?

Bioreactors are used for large-scale cell culture to produce proteins, often in eukaryotic cells, like mammalian cells, which facilitate proper protein folding and modifications.

How does the lac operon system control gene expression in plasmid vectors?

The lac operon uses a repressor protein that binds to the operator to prevent transcription. An inducer molecule can inhibit the repressor, allowing gene expression to occur.

What is blue-white screening, and how is it used in recombinant plasmid identification?

Blue-white screening identifies recombinant plasmids by disrupting the lacZ gene. White colonies indicate successful gene insertion, as the lacZ gene's disruption prevents beta-galactosidase production, which would turn colonies blue.