L19: Recombinant DNA Technology

Question 1

Why was treatment in the 1950s for diabetes (DNA sequencing) not always 100% pure?

Answer 1

Because insulin from cattle and pigs were used so the DNA was not entirely the same.

Question 2

This is joining bits of DNA together (sometimes from different species). These are then inserted into a organism to produce a useful protein.

Answer 2

Recombinant DNA technologies

Question 3

These are circular pieces of double stranded DNA. They are the critical elements of recombinant DNA technologies.

Answer 3

Plasmids

Question 4

True or false. Plasmids are only found in some eukaryotes and all prokaryotes.

Answer 4

False.

Question 5

True or false. Plasmids provide antibiotic resistance to hosts.

Answer 5

True (which gives selective advantage for bacteria)

Question 6

This is a component of the plasmid that allows initiation of replication using host, DNA polymerase.

Answer 6

Origin of replication (Ori)

Question 7

This is a component of the plasmid that allows selection of cells containing plasmid.

Answer 7

Antibiotic resistance gene

Question 8

This is a component of the plasmid that drives the expression of your favourite gene in cells with appropriate transcription factor machinery. (These have to be specific to the gene/organism)

Answer 8

Promoter (animal-specific)

Question 9

Why are there different promoters of the same gene in different organisms?

Answer 9

Because it is expressed in different cell types (like, the same gene but in a fish cell, or the same gene but in a neuron)

• it is just because the gene is expressed in a different cell type.

Question 10

What are the 3 key components of plasmids?

Answer 10

Origin of replication, promoter, and the antibiotic resistance gene

Question 11

These are responsible for cutting dsDNA (double stranded) at specific sequences.

Answer 11

Restriction enzymes

Question 12

True or false. Bacteria do not have restriction enzymes, only humans.

Answer 12

False. Restriction enzymes are naturally found in bacteria which makes them such good plasmids.

Question 13

What other function do restriction enzymes have in bacteria?

Answer 13

They provide defense system to degrade foreign DNA.

Question 14

This is responsible for catalysing the formation of phosphodiester bond to repair nick in DNA backbone. It has complementary base pairing functions.

Answer 14

DNA ligase

Question 15

This is the process of transferring plasmids into bacteria. recap: what’s a plasmid?

Answer 15

Transformation

A plasmid is a circular double stranded piece of DNA

Question 16

These can replicate independently of the host’s chromosomal DNA.

Answer 16

Plasmids.

Question 17

Outline the process of amplifying plasmids.

Answer 17

1) Transformed bacteria selected by antibiotic resistance gene.

2) Expression of plasmid gene in bacteria (through bacterial promoter)

3) Amplification of bacteria and purification of DNA (origin of replication)

Question 18

What does the universal genetic code mean?

Answer 18

All organisms read the same codons as the same amino acids.

Question 19

What is the significance of the genetic code being universal?

Answer 19

It means we can transform a human gene into bacteria and make it produce the same protein.

How can it produce the same protein? Because bacterial machinery can read the same codons and express them as the same proteins (as is in humans)

Question 20

What is the main problem with human genes/proteins being processed in prokaryotes/bacteria?

Answer 20

Prokaryotes don’t have introns- so do not have the machinery to splice them out of the coding sequence.

Question 21

True or false. A universal genetic code enables only some genes from one species to be expressed in another species.

Answer 21

False.