Plasmids

Question 1

What are some of the uses for genetic transformation?

Answer 1

• agriculture: coding for resistance against frost, pest, or spoilage
• Bioremediation: bacteria can be genetically transformed with genes enabling them to digest oil spills
• medicine: diseases caused by defective genes can be treated by gene therapy

Question 2

T/F: all plasmids are vectors

Answer 2

false

Question 3

What is a plasmid?

Answer 3

small, circular, double stranded, extrachromasomal elements found in some bacteria strains

Question 4

since plasmids are closed circular double-stranded DNA molecules they are under…?

Answer 4

super helical tension and negative supercoiling

Question 5

T/F: all plasmid forms have the same molecular weight.

Answer 5

true

Question 6

What genes are encoded on plasmid DNA?

Answer 6

• genes for its own replication and propogation
• origins of replication
• gene for partitioning plasmids into each daughter cell during cell division
• genes that give advantages to their bacterial hosts.

Question 7

what is a common plasmid advantage?

Answer 7

ampicillin resistance

Question 8

What is a selectable marker?

Answer 8

A plasmid is coded for a specific thing ie antibiotic resistance and will kill off everything except that speicfic thing

Question 9

what is a multiple cloning site?

Answer 9

requires the removal and addition of various restriction enzyme sites so a region on the plasmid would contain a number of unique restriction enzyme sites

Question 10

what are regulatory signals

Answer 10

regulates expression of cloned genes

Question 11

how do promoter sequences help plasmids?

Answer 11

allows expression (transcription) of gene cloned into plasmid

Question 12

what is a pGLO plasmid?

Answer 12

green fluorescent protein gene in jellyfish

Question 13

what is pGLO induced by?

Answer 13

the sugar arabinose

Question 14

what is pGLO for?

Answer 14

ampicillin resistance

Question 15

what is the process of gene cloning in bacteria?

Answer 15

• isolates the fragment of DNA containing the gene of interest
• fragment is inserted containing the gene to produce recombinant DNA molecule
• then introduce the recombinant DNA into host cell
• plate transformed bacteria so it can grow and form a colony
• colony then produces many new recombinant bacteria plasmids

Question 16

how many times should a plasmid be cut?

Answer 16

1-2 since more will shred the DNA (even 2 is a lot)

Question 17

What is transformation of plasmids?

Answer 17

insertion of free DNA into the cell

Question 18

What is transduction of plasmids?

Answer 18

DNA containing virus is inserted into the bacteria cell

Question 19

What is conjugation of plasmids?

Answer 19

plasmid is placed into donor cell which is then transferred into a secondary cell

Question 20

what is the specialized limb in the conjugation of plasmids?

Answer 20

conjugation tube

Question 21

What is the hypothesized reason for CaCl2 treatment?

Answer 21

creating pores and priming the cells

Question 22

what is the heat-shock method?

Answer 22

incubate bacterial cells with cold solution of CaCl2 which makes pores adding the DNA to the cells and then the incubation heat shocks it

Question 23

explain electroporation:

Answer 23

short current pulses create pores and the bacteria is washed with cold H2O to remove residual ions , DNA is added

Question 24

what is lipofection?

Answer 24

DNA is encapsulated by a micelle and fused to a cell membrane

Question 25

what is the issue with lipofection?

Answer 25

it is expensive

Question 26

What are the four methods of introducing DNA into bacteria?

Answer 26

1. microinjection
2. electroporation
3. lipofection
4. ballistic/gene gun