Lab 5

Question 1

Where do restriction enzymes cut?

Answer 1

At the DNA backbone.

Question 2

Restriction enzyme site:

Answer 2

The target site from which the DNA of interest is attached to the plasmid.

Question 3

Plasmid:

Answer 3

Extra-chromosomal DNA that replicates independently of a chromosome.

Question 4

A recombinant plasmid:

Answer 4

A plasmid that contains genes from two different sources.

Question 5

DNA ligase:

Answer 5

Glues two pieces of DNA together.

It uses Phosphodiester bonds to connect the backbones together and ATP is used as an energy source.

Question 6

Another name for a plasmid:

Answer 6

Vector.

Question 7

What does a vector not have:

Answer 7

An insert.

Question 8

Most plasmids contain:

Answer 8

Antibiotic resistance gene.

Question 9

Plasmids’ ORI:

Answer 9

Origin of replication.
This is needed for the plasmid to be replicated in the bacterium.
ORI is where replication begins.

Question 10

Polylinker / Polycloning:

Answer 10

Contains several restriction enzyme sites that are very close together.

This is where the gene is inserted.

Question 11

Bacteria Transformation:

Answer 11

The process of moving a plasmid into a bacteria.

Question 12

pHUG21 contains:

Answer 12

A heme transport gene.
An ampicillin resistance gene.

Question 13

Another name for restriction enzyme:

Answer 13

Restriction endonuclease.

Question 14

Restriction endonuclease:

Answer 14

An enzyme that cuts the DNA at a specific DNA sequence within the DNA strands, rather than at the very end.

Question 15

Types of ends a restriction enzyme can leave:

Answer 15

Sticky end [Cohesive/overhang]
Blunt end.

Question 16

Another name for sticky end:

Answer 16

Cohesive end.
Overhang.

Question 17

Palindromic sequence:

Answer 17

A sequence that reads the same backward and forwards.

Question 18

Restriction enzymes cut at:

Answer 18

Specific sites.

Question 19

Restriction enzymes recognize:

Answer 19

The entire sequence.

Question 20

Blunt end:

Answer 20

No overhang, harder to hydrogen bond.

Question 21

How many more pieces of inserts than plasmids do you need?

Answer 21

4 times as many.

Question 22

Why do bacteria make restriction endonuclease:

Answer 22

Bacteria produce them to chop up DNA from a bacteriophage that may have infected the bacterium.

Question 23

The plasmid and the DNA need to have the same:

Answer 23

Overhangs.

Question 24

Methylase:

Answer 24

Methylate some of the bases in the sequence so the restriction enzyme cannot cut it.

Question 25

Restriction enzyme modification:

Answer 25

Made up of the restriction enzyme and the methylase.

Question 26

Why is modification done:

Answer 26

To prevent degradation of the host DNA by its restriction endonuclease.

Question 27

Where is methylation done:

Answer 27

On the bases.

Question 28

Restriction endonuclease nomenclature
Ex: E. coli RY13
1.) The first letter represents:
2.) The 2nd and 3rd letters represents:
3.) The second capital letter represents:
4.) The number represents:

Answer 28

EcoR1
1.) The genus.
2.) Species the enzyme was isolated.
3.) The strain.
4.) The order in which the enzyme was
discovered.

Question 29

How do molecular biologists use restriction enzymes:

Answer 29

They use them to cut up DNA of interest into smaller pieces which are easier to manipulate.

Question 30

Restriction enzyme cuts:

Answer 30

On both sides of the gene of interest.

Question 31

kB stands for:

Answer 31

Kilobases.

Question 32

Recombinant plasmids can be used by researchers to:

Answer 32

Overexpress a gene so protein can be purified and studied more closely.

Question 33

Expression vectors:

Answer 33

Vectors that are used to overexpress a gene.

Question 34

Restriction enzyme map can:

Answer 34

Help you create a plasmid that contains only a certain gene.