Question

A) Describe the differences between expressed sequences and not expressed DNA sequences and how to study them. For example, cDNA vs genomic libraries and RT PCR vs PCR-

B) Restriction cutting DNA, running gels, and ligating DNA to make recombinant DNA.

C) Difference between southern, northern, and western blotting. Describe what each is looking for?

Answer 1

A. Expressed sequences produce RNA and protein. They are in the euchromatic state. We can study the expression level of a sequence by quantifying the transcript (qRT PCR) or by quantifying the protein (Western blotting).

cDNA libraries contain only the expressed portions of the genome whereas genomic libraries contain all the sequences of the genome.

RT-PCR is performed to obtain cDNA from mRNA. PCR is used to amplify DNA sequences from DNA template.

B. Restriction endonucleases are enzymes that recognize specific sequences and cleave. They are very useful in gene cloning.

See image.

C. Southern blotting is used to detect DNA sequences using complementary DNA probes.

Northern blotting is used to detect RNA sequences using complementary DNA/RNA probes.

Western blotting is used to detect proteins using antibodies.