A2 functional diversity through mRNA splicing

Question 1

how many introns does the average human gene have and what size are they

Answer 1

8 but some have >100
from 50 to 10,000 nts

Question 2

what does the cap on pre-mRNA involve

Answer 2

involves methylation of bases
and formation of rare 5’ to 5’ triphosphate bridge with GTP
occurs when pre-mRNA is around 25 nt long
involves specific capping proteins
enhances stability and is key for efficient translation

Question 3

what does the Poly(A) tail on pre-mRNA involve

Answer 3

Poly A added by enzyme called poly(A) polymerase
(A)n where n may be 250+
increases stability and enhances translation

Question 4

what do all introns start and end with

Answer 4

start with GU and with AG

Question 5

what is the key to splicing introns

Answer 5

a pyrimidine rich tract (PY)n and a specific adenine

Question 6

splicing proteins recognise specific sites, what are these

Answer 6

the GU at the 5’ splice site and the branch site

Question 7

the proteins involved in splicing recruit other proteins into the large complex called what

Answer 7

spliceosome

Question 8

what can the spliceosome do

Answer 8

can capture, splice, and release RNA accurately and in a coordinated way which involves a careful choreography of spliceosome components

Question 9

what is alternative splicing

Answer 9

a process whereby different mRNAs are generated from the same initial (primary) transcript

Question 10

what can cause disease by affecting splicing

Answer 10

mutations

Question 11

what can happen when splicing goes awry

Answer 11

formation of a non functional protein or mutated one

Question 12

how can some RNA mols self splice

Answer 12

because RNA can fold into distinct structures it has the capacity to act as an enzyme and splice itself

Question 13

what drives splicing within the RNA structure

Answer 13

short stretches of nts on RNA can bp with other regions creating folds which in turn drives splicing

Question 14

who discovered that RNA can act as an enzyme and self splice

Answer 14

tom cech and sidney altman

Question 15

what is RNA interference

Answer 15

RNAi is a powerful tool to disrupt gene expression
discovered when double stranded RNA was introduced into a cell
found to suppress the transcription of genes that contained sequences present in the original double stranded RNA

Question 16

what does RNAi reply on

Answer 16

key activity of small interfering RNAs (or siRNAs)

Question 17

what is RNAi widely used in SIPBS for

Answer 17

to silence the expression of specific genes

Question 18

why is RNAi exploited in cell biology

Answer 18

to examine the function of a particular gene product by silencing other genes

Question 19

what is the name given to genome editing

Answer 19

CRISPR/Cas

Question 20

technology behind CRISPR/CAs

Answer 20

specific guide gRNA is synthesised that directs an endonuclease enzyme called Cas( to specific DNA target sequence
can be used to create a cleavage in the DNA at a precise point and so insert or delete nucleotides (or create a frameshift)

Question 21

what are the potential uses or CRISPR

Answer 21

-poweful lab tool to create cell lines to probe specific gene function by creating a ‘knockout’
-can produce specific mutations in the germ line- probe function of key putative residues in a protein
-can be used ex vivo and in vivo genome editing for clinical therapy

Question 22

ex vivo genome editing for clinical therapy

Answer 22

cells isolated from a patient to be treated
edited then regrafted back to patient
to achieve therapeutic success, the target cells must be able to survive in vitro ad=nd return to the target tissue after transplantation

Question 23

in vivo genome editing for clinical therapy

Answer 23

engineered nucleases are delivered by viral or nonviral approached and directly injected into the patient for systemic or targeted tissues (ie eye, brain, or muscle) effect