Lecture 2

Question 1

What are some vector types?

Answer 1

Plasmid vector, Lamda phage vector (larger DNA sequences), Cosmids (packaged into phage for transmission and use plasmid genes to replicate in host cell), Expression vectors.

Question 2

What are markers and why are they used?

Answer 2

Selectable marker (e.g Antibiotic resistance genes) are genes with specific qualities that exist or are inserted into the vector. They aid in seeing which bacteria has successfully taken up plasmid as when grown on a specific media, the bacteria with the plasmid with the marker will be able to survive

Question 3

What is transformation?

Answer 3

The process where the bacteria cell takes up the plasmid

Question 4

How is restriction enzyme used to make recombinant DNA?

Answer 4

Restriction enzyme cuts the vector and fragment of DNA at the same palindromic sequence
DNA ligase then seals the two strands together, forming the recombinant plasmid

Question 5

…. and …. showed that e.coli treated with CaCl2 could take up DNA and … showed it could also take up a plasmid

Answer 5

Mandel and Higa showed that e.coli treated with CaCl2 could take up DNA and cohen showed it could also take up a plasmid

Question 6

What is the process of bacterial transformation

Answer 6

1) Bacteria culture is centrifuged and chilled in ice generating competent cells stored at -80C
2) Plasmid DNA added ans stored in ice for half an hour
3) Put in a water bath qat 42 degrees (heat shock)
4) put back onto liquid medium (1hr) plate out onto surface medium

Question 7

Electroporation

Answer 7

Small electrical current causes bacterial membrane to become permeable so plasmid can be taken up