chapter 14: site specific recombination-- exam 2

Question 1

what is site specific recombination?

Answer 1

rearrangement between 2 specific DNA sequences

Question 2

What does site specific combination result in?

Answer 2

• insertion
• deletion
• inversion

Question 3

Site specific recombination requires ______ and ______

Answer 3

• restriction endonucleases
  and
• ligases

Question 4

T/f Site specific recombination is isoenergenic and doesn’t require extensive homology

Answer 4

true

Question 5

What is the earliest form of parasitism?

Answer 5

Transposition

Question 6

What is transposition?

Answer 6

When transposons move freely between different chromosomes

Question 7

Where are recombination sites located?

Answer 7

at inverted repeats with a non palindromic core

Question 8

Does the non palindromic core have direction?

Answer 8

yes

Question 9

Describe the direction of the non palindromic core

Answer 9

• recombination with cores in opposite directions lead to inversions
• recombination with cores in the same directions lead to deletions
• between 2 separate chromosomes, same direction cores lead to insertions

Question 10

recombination with cores in opposite directions lead to ______

Answer 10

inversions

Question 11

recombination with cores in the same directions lead to _______

Answer 11

deletions

Question 12

between __________, same direction cores lead to ______

Answer 12

2 separate chromosomes ; insertions

Question 13

Active sites are typically ___ or ___ residues as nucleophile

Answer 13

Ser ; Tyr

Question 14

_ light gray recombinases are active at a time

Answer 14

2

Question 15

site specific reaction is very similar to the process of _____

Answer 15

topoisomerases

Question 16

What are some examples of site-specific recombination?

Answer 16

• plasmid replication
• viral DNA circularization
• biotech
• phase variation

Question 17

What are bacteriophages?

Answer 17

viruses that infect bacteria

Question 18

What are the 2 pathways of viral infections?

Answer 18

1. lysogenic pathway

2. lytic pathway

Question 19

LoxP is…

Answer 19

site specific recombination loci

- allows for circulation of bacteriophage DNA

Question 20

What is Salmonella typhimurium?

Answer 20

Regulates gene expression via site-specific recombination
•Flagella is a prominent target of mammalian immune systems
•Can change protein types of flagella to avoid host immunity!
•FljB  FliC

Question 21

Pieces of DNA that move from one part of the genome to the other (donor –> target site)
- can be random or specific

Answer 21

Transposons

Question 22

What are the 3 types of transposons?

Answer 22

1. cut and paste
2. replicative
3. RNA intermediate (retrotransposon)

Question 23

________ is coded for within the transposable element transposon

Answer 23

transposase

Question 24

What 2 things does transposes catalyze?

Answer 24

1. hydrolytic cleavage

2. Transesterification

Question 25

What is a cut and paste transposition?

Answer 25

•Double-stranded cuts in donor DNA—must be repaired* •Liberated DNA attacks target DNA via 3’ –OH groups to form new phosphodiester bonds •Single-stranded gaps filled in with repetitive DNA *Donor DNA can be repaired through ligation or recombination repair

Question 26

What are the 2 types of retrotransposons?

Answer 26

1. Long-terminal repeat with extrachromosomal primed retrotransposition (LTR (EP)) 2. Non-long-terminal repeat with target-primed retrotransposition (Non LTR (TP))

Question 27

LTR (EP) Retrotransposons:

Answer 27

* Transposon with a long-terminal repeat and extrachromosomally primed retrotransposition * Long-terminal repeat = repetitive DNA at the termini of transposon between 100 – 5000 bp * Extrachromsomally primed = DNA primers come from tRNA (not part of the chromosome)

Question 28

Mechanism of LTR (EP) Retrotransposons:

Answer 28

1. Transposon is transcribed by RNA polymerase  ssRNA transcript 2. Reverse transcriptase  ds-cDNA * •Remember, DNA always requires a primer! In this case . . . tRNA . . . wild! 3. Circularized by integrase (related to transposase) 4. Invasion of target DNA 5. Donor and target both have copy of transposon * Reverse transcriptase can make dsDNA out of ssRNA. That is, it can use DNA and RNA as a template.

Question 29

Non-LTR (TP) Retrotransposons

Answer 29

* Transposon without long-terminal repeat, with target-primed retrotransposition * Uses part of the ssRNA intermediate as a primer for DNA

Question 30

Bacteria have 3 common classes of transposons, what are they?

Answer 30

1. Insertion sequences 2. Composite transposons 3. Complex transposons

Question 31

Generally, do bacterial transposons have RNA intermediates?

Answer 31

NO

Question 32

What do Insertion sequences contain?

Answer 32

Only DNA sequences necessary for transposition

Question 33

What do composite transposons contain?

Answer 33

DNA sequences necessary for transposition and other genes

Question 34

Complex transposons have:

Answer 34

- large genome - genes for other processes than transposition - act as transposon or bacteriophage DNA (lytic and lysogenic pathway_

Question 35

What percent of human DNA in each cell consists of transposons?

Answer 35

46%

Question 36

What percent of transposons in human DNA come from retrotransposons?

Answer 36

90%

Question 37

What percent of humans retrotransposons in their DNA are LTR vs Non-LTR?

Answer 37

LTR: 20% | Non- LTR: 80%

Question 38

Describe LTR retrotransposons:

Answer 38

* Similar to viruses * Transcribed with a poly-A tail•Transported to cytoplasm * Translated to * Gag = structural; creation of VLP * Pol = reverse transcriptase * Int = integrase * Reverse transcribed and brought into the nucleus via VLP * dsDNA integrates in new location

Question 39

What are the 2 types of Non-LTR transposons?

Answer 39

LINEs and SINEs

Question 40

What does LINEs stand for?

Answer 40

Long interspersed nuclear elements

Question 41

What does SINEs stand for?

Answer 41

Short interspersed nuclear elements

Question 42

Are LINEs autonomous?

Answer 42

Yes

Question 43

Are SINEs autonomous?

Answer 43

No

Question 44

Describe LINEs:

Answer 44

* Transcribed to RNA in nucleus * RNA translated into cytoplasm * Reintegrates DNA into the nucleus * Autonomous

Question 45

Describe SINEs:

Answer 45

•Transcribed to RNA in nucleus •RNA translated into cytoplasm •Reintegrates DNA into the nucleus - Non-autonomous

Question 46

What is the difference between SINEs and LINEs?

Answer 46

SINEs are non-autonomous

Question 47

What are retroviruses?

Answer 47

Viruses that contain reverse transcriptase

Question 48

What are the typical genes of retroviruses?

Answer 48

``` Gag = structural Pol = reverse transcriptase, integrase, proteases Env= structural ; allows element to transfer from cell to cell ```

Question 49

What does viral reverse transcriptase lack?

Answer 49

3' --> 5' exonuclease activity

Question 50

HIV is a retrovirus that infects ____ cells

Answer 50

T

Question 51

What are the drug targets for HIV

Answer 51

- reverse transcriptase | - HIV proteases

Question 52

Why is combination effective in fighting HIV?

Answer 52

because there is a very high mutation rate