Part D: Lecture 33

Question 1

Recombinant chromosome made in ___

Answer 1

in vivo by crossovers

Question 2

recombinant DNA made in ___

Answer 2

in vitro by geneticists (DNA from two organisms)

Question 3

Cloning (def.)

Answer 3

making identical copies of something

Question 4

gene cloning (def.)

Answer 4

using bacterial cells to make identical copies of a specific gene

Question 5

Tools needed for gene cloning

Answer 5

-insert DNA + vector DNA -> (ligate) recombinant DNA
–> (transform bacteria cell)

Question 6

plasmids need _____ to be reproduced in bacterial cells

Answer 6

ori (origin)

Question 7

bacteriocide gene do what?

Answer 7

cells that have it kills cells that don’t have it

Question 8

restriction enzymes cut where on DNA?

Answer 8

the sugar-phosphate backbone

Question 9

T4 ligase from bacteriophage does what?

Answer 9

reforms sugar-phosphate backbone (covalent bonds)

Question 10

T4 ligase does not do what?

Answer 10

form base pairs, base pairs are formed by spont. H bonds

Question 11

Lab strains are used in transforming bacterial cells because ____

Answer 11

-they won’t digest your DNA (have mutations in genes for restriction enzymes)
-can’t escape the lab (would die in the wild)- auxotrophic
-plasmids can enter (holes in cell cell)

Question 12

Transformation procedure

Answer 12

Step 1 - Prepare the insert DNA (digestion with RE)
Optional Step - Gel purify the insert DNA
Step 2 - Prepare the vector DNA (digestion with RE)
Step 3 - Mix the DNAs and add T4 DNA Ligase
Step 4 - Transform competent E. coli cells (1 in 1M)
Step 5 - Plate cells (cell with plasmid survive)
Step 6 - Remove cells from the colony (save: -70C & test)
Test: recombinant plasmid + RE + AGE

Question 13

AGE stands for ____

Answer 13

Agarose Gel electrophoresis