Chapter 9: Biotechniology and recombinant DNA

Question 1

What are the steps involved in cloning a gene?

Answer 1

A vector, such as a plasmid is isolated. DNA containing the gene of interest from a different species cleaved by specific restriction enzymes. The desired gene is selected and inserted into the plasmid. The plasmid is taken up by a cell such as a bacterium. Cells containing the plasmid with the gene of interest with either of two goals in mind: to create and harvest copies of a gene or create and harvest products of a gene.

Question 2

What are the 5 ways to get foreign DNA into cells?

Answer 2

Transformation, electroporation, protoplasts, protoplast fusion, and microinjection.

Question 3

Transformation

Answer 3

cells take up DNA from the surrounding environment. Can be treated with solutions and heat shocked to be more likely to take up the DNA.

Question 4

Electroporation

Answer 4

uses electrical current to form microscopic pores in the membranes of cells, and DNA enters through the pores

Question 5

Protoplasts

Answer 5

produced by enzymatically removing the cell wall, thereby allowing more direct access to the plasma membrane

Question 6

Protoplast fusion

Answer 6

protoplasts in solution fuse slowly, but with the addition of polyethylene glycol, the frequency of fusion goes up. In the new hybrid cell, the DNA derived from the two “parent” cells may undergo natural recombination.

Question 7

Microinjection

Answer 7

micropipette punctures the cell membrane and DNA can be injected through it.

Question 8

Briefly describe PCR

Answer 8

DNA Incubate 94C to separate strands of DNA; Primers attach to the single stranded Target DNA during incubation at 60C; Solution is incubated at 72* C; Elongation happens during the incubation period as the DNA polymerase copies the Target DNA.

Question 9

Applications of Recombinant DNA technology

Answer 9

1. Therapeutic applications: producing pharmaceuticals, vaccines, and gene therapy. 2. Scientific applications: DNA sequencing (genome projects), forensics and fingerprinting. 3: agricultural applications - Altering nitrogen fixing capabilities of plants (altering host specificity for rhizobium) and engineering plants to have natural pesticide (Bt toxin from Bacillus thuringiensis)