CRISPR - MT3 - Part 1

Question 1

What is CRISPR? (2)

Answer 1

1. It is a technique for finding any place on any genome you wish to find and making any alteration you wish to make
2. It is a gene locus in many bacteria
   - it is a certain place on the bacterial chromosome that codes for certain proteins and/or has certain functions other than coding

Question 2

What does CRISPR stand for?

Answer 2

Clustered Regularly Interspaced Short Palindromic Repeats

Question 3

Locus

Answer 3

Is a fixed position on a chromosome, like the position of a gene or a marker

Question 4

What is CRISPR the latest example of?

Answer 4

A molecular tool in biology that is derived from a natural structure or process

Question 5

Plasmid

Answer 5

A genetic structure is a cell that can replicate independently of a chromosome
- easy to manipulate in a test tube

Question 6

Viruses

Answer 6

An infective agent that consists of nucleic acid molecules in a protein coat, is too small to be seen by light microscopy and is bale to multiply only within the living cells of a host

Question 7

Restriction enzyme

Answer 7

An enzyme produced by bacteria, having the property of cleaving DNA molecules at or near a specific sequence of bases

Question 8

DNA ligase

Answer 8

Is a specific type of enzyme, it facilitates the joining of DNA strands together by catalyzing the formation of a phospho-diester

Question 9

DNA polymerase

Answer 9

Is a type of enzyme that is responsible for forming new copies of DNA, in the form of nucleic acid molecules

Question 10

What is an example of DNA polymerase?

Answer 10

Taq polymerase

Question 11

What 3 things are needed when cloning a gene?

Answer 11

1. Restriction enzymes
2. DNA ligase
3. Plasmid vector

Question 12

What can bacteria be infected with?

Answer 12

Viruses (phages)

Question 13

What have bacteria evolved?

Answer 13

Defences against viruses

Question 14

What do bacteria have on them?

Answer 14

A selective pressure on them to develop a defensive mechanism

Question 15

What do phages encode?

Answer 15

Some functions needed for replication

- a typical virus will make a few of these proteins and then hijack the cell

Question 16

What do viruses not have?

Answer 16

A selective barrier

Question 17

What is one kind of innate defence system in bacteria?

Answer 17

Restriction enzymes

- non-acquired, non-specific

Question 18

What does methylation do?

Answer 18

Prevents cutting of the DNA of the attacked cell

- restriction enzymes font cut the bacterial DNA because they are methylated

Question 19

What is one kind of adaptive defence system?

Answer 19

CRISPR

• acquired, specific
• changes to individual experience

Question 20

What happens in the first exposure to a virus?

Answer 20

A bacterial cell can chop up the viral genome into small fragments and insert some of the fragments into the bacterial CRISPR locus

Question 21

What must the viral genomic fragments be next to?

Answer 21

The sequence 5’-NGG-3’ in the viral genome

- this is the PAM

Question 22

PAM

Answer 22

Protospacer adjacent motif

Question 23

What does the CRISPR locus consist of? (2)

Answer 23

1. A number of short palindromic repeats
   - the viral fragments are interested as non-repetitive spacers between the repeats
2. Contains genes such as cas or CRISPR-associated genes that encode for proteins, such as nucleases that can cut DNA

Question 24

What are new viral fragments interested in?

Answer 24

The closest available position to the leader sequence

Question 25

What do viral sequence represent?

Answer 25

Molecular memories of the viruses they have encountered, just our own memory lymphocytes encodes these memories in us

Question 26

What happens if the cell encounters the same virus again? (2)

Answer 26

1. The CRISPR locus is transcribed to CRISPR derived RNA (crRNA)
2. The nearby cas genes are expressed

Question 27

How can viral DNA find the same sequence in the invading viral genome?

Answer 27

By homologous base pairing