Chapter 2

Question 1

Fertility (F) plasmids

Answer 1

Allow for conjugation and transfer of DNA between bacterial cells

Question 2

Resistance plasmids

Answer 2

contain genes that encode for resistance to drugs or antibiotics

Question 3

Virulence plasmids

Answer 3

contain genes that can cause a bacterium to behave pathogenic

Question 4

Plasmid

Answer 4

A small circular extra chromosomal DNA that can be replicated autonomously in the cell

Question 5

Cloning vector

Answer 5

Plasmids or DNA that has been engineered to allow for foreign DNA to be inserted and propagated

Question 6

pBR332

Answer 6

Created
by
Bolivar
and
Rodriguez
in
 1977.

Thought 
to 
be
 the 
first 
(or 
at 
least
 one 
of 
the 
first)
 vectors
designed
 to
 clone 
(insert) DNA
–
a 
key 
in
 recombinant 
DNA
technology.


Has 
the 
ability 
to 
replicate 
in
 bacterial
 cells 
independently
 of
 chromosomal 
replication.


Contains 
resistance 
genes 
to
 ampicillin
 (amp
R) 
and
tetracycline
 (tet
R).


Has 
over
 40
 sites 
recognized
 by
 restriction 
enzymes 
that
 only 
cut
 the
 vector 
once.


Question 7

3 Features required of cloning vectors

Answer 7

1) Replication origin - a sequence that is necessary for the initiation of DNA replication
2) Selectable marker - a gene that confers the ability to distinguish bacteria that contain the plasmid from bacteria that do not
3) Unique restriction sites - vector mus have sites where it can be cut only once to insert DNA

Question 8

Bacteriocidal

Answer 8

Kills the bacteria that are not resistant

Ex = Ampicillin, Kanamycin, Streptomycin

Question 9

Bacteriostatic

Answer 9

Prevents growth of bacterial cells that are not resistant

Ex = Tetracycline, Chloramphenicol

Question 10

Multiple cloning site (MCS)

Answer 10

a region that contains a number of sites in a small region that are unique to that particular vector

Question 11

Selection

Answer 11

Only the cells you want can survive and grow; anything else does not grow or is killed

Question 12

Screen

Answer 12

All cells grow; however, the desired cells are easily distinguished form the ones you don’t want

Question 13

Screen by resistance

Answer 13

Plasmid is Amp and Tet resistant. Cut Tet with BamHI. Insert DNA.
Select for cells that confer ampicillin resistance (contain plasmid). Screen for cells that do not confer tetracycline resistance (contain plasmid with inserted DNA)

Question 14

Screen by blue white color

Answer 14

lacZ’ gene produces β-galactosidase which breaks down X-gal to produce a blue color. Cut lacZ’ gene and insert new DNA

1) Select for cells resistant to ampicillin (contain plasmid)
2) Screen for cells that are white in color when plated on media containing X-gal( contain plasmid with inserted DNA)

Question 15

Plasmid

Answer 15

A cloning vector with a bacterial replication origin, selectable marker, and unique restriction sites. Can insert DNA up to about 20 kbp in lenght

Question 16

Shuttle vector

Answer 16

a cloning vector that can be inserted into two different organisms. Has similar features to the standard cloning plasmid

Question 17

Bacteriophage

Answer 17

a bacterial virus; when manipulated for cloning DNA, can accommodate DNA fragments up to about 25 kbp in length

Question 18

Cosmid

Answer 18

a cloning vector that can be packaged into a bacteriophage λ, such that inserted DNA can be up to about 45 kbp in length

Question 19

Bacterial artificial chromosome (BAC)

Answer 19

A cloning vector that is based off of the fertility plasmid and has replication and partitioning functions allowing for stable heritability. Can accommodate DNA up to about 300 kbp in lenght

Question 20

Yeast artificail chromosome (YAC)

Answer 20

contains replication origin, centromere, telomeres, and a selectable marker. Can accommodate DNA up to about 1 Mbp in lenght

Question 21

Auxotrophic

Answer 21

when a cell requires a compound (i.e. an amino acid) because it is unable to synthesize this compound itself. Usually due to a mutation in a bio synthetic pathway

Question 22

Prototrophic

Answer 22

when a cell does NOT require a compound in the media because it is able to synthesize this compound itself

Question 23

Yeast episomal plasmid

Answer 23

a shuttle vector that autonomously replicates yeast due to the 2μ DNA (containing a replication origin). A high copy number plasmid that is often used to over express genes in yeast

Question 24

Yeast integrating plasmid

Answer 24

a shuttle vector that does not autonomously replicate (lacks replication origin). Only way to be replicated is to integrate into the genome

Question 25

Yeast centromeric plasmid

Answer 25

a shuttle vector that autonomously replicates and contains a centromere. A low copy number plasmid that is very stably inherited due to the centromere which allows for proper segregation of the plasmid during mitosis and subsequent cell division