Recombinant DNA and gene projects

Question 1

How is a copy of a gene of interest got from DNA code

Answer 1

gene probe identifies DNA
restriction enzymes cut the DNA
forming sticky ends

+forms sticky ends
-contains introns, requires base sequencing

Question 2

How is a copy of a gene of interest got from mRNA

Answer 2

section on DNA is identified
cell transcribes and splices mRNA
mRNA isolated and added to reverse transcriptase

+no sequencing needed, no introns
-doesn’t form sticky ends

Question 3

How is a copy of a gene of interest got from the gene machine

Answer 3

sequence types into the automated polynucleotide sequencer
fragment created

+no introns, section found quickly
-no sticky ends, base sequence needed

Question 4

What are the three stages of in vivo cloning (PCR)

Answer 4

denaturation
annealing
extention

Question 5

What happens in denaturation

Answer 5

PCR machine heats to 95
hydrogen bonds break
two strands of DNA separate

Question 6

What happens during annealing

Answer 6

PCR machine cools down to 55-65
primer sequences complementary base pair to the fragments

Question 7

What happens during extension

Answer 7

PCR machine heats to 72
primers are extended to form complementary DNA
free floating nucleotides complementary base pair to the DNA
phosphodiester bonds formed by taq polymerase

Question 8

Heat shock

Answer 8

-mix bacteria with plasmids
-add calcium salts to stop repulsion
-freeze
-rapidly increase temp to 40 to create pores in the bacterial cell membrane which allows plasmids to pass through

Question 9

Electroporation

Answer 9

-electric charge passes through phospholipid bilayer
-hydrophobic tails repelled
-tails turn away from charge to pores in the membrane form

Question 10

Forming a transgenic cell

Answer 10

-cut out bacterial plasmid with restriction enzymes to produce sticky ends
- DNA fragment cute to have complementary sticky ends form hydrogen bonds with comp bases
-DNA ligase joins bacterial plasmid and DNA fragment, forming phosphodiester bonds