Zaidi Analyzing Cells/Systems 2

Question 1

Restriction Endonucleases

Answer 1

Enzymes isolated form bacteria - cut DNA at specific sites depending on the type of endonuclease

Question 2

Gluing DNA together aka the ___ rxn is easier with compatible cohesive ends.

Answer 2

ligase

Question 3

To put a DNA fragment into a plasmid, making recombinant DNA, you need to use the ___ ___ to cut the original DNA.

Answer 3

restriction nuclease.

Question 4

cDNAs are what? What does a cDNA require?

Answer 4

DNA copy of mRNA, contains no introns.

Requires Reverse Transcriptase.

Question 5

There are two different libraries used to store genes. What are they?

Answer 5

Genomic DNA and cDNA library.

Question 6

What ingredients are necessary for the PCR cycle?

Answer 6

DNA sample
Primers (oligos)
Heat-stable DNA polymerase.
Deoxyribonucleotides
Thermocycler.

Question 7

What are the three steps for PCR?

What is the purpose of PCR?

Each cycle increases the amount of DNA molecules this many times?

Answer 7

1. Heat to separate (denature) strands into single stranded DNA.
2. Cool to anneal primers
3. DNA synthesis.

To amplify any DNA in the sample.

Each cycle doubles the previous amount of DNA.

Question 8

Primers flank each end of DNA in what direction. The primers __ and these, ___, molecules are added one by one.

What is the purpose of Taq Polymerase?

Answer 8

3’ to 5’

dNTPs (A/T/G/C).

Taq polymerase synthesizes copy of DNA by extending the primers on both ends.

Question 9

List Advantage and Disadvantage of PCR:

Answer 9

Advantage: very small amount of DNA is needed.

Disadvantage: Need to know the sequence of the flanking DNA primer.