nucleic acid part 4

Question 1

what was the aim of the human genome project ?

Answer 1

to determine the human genome sequence , this would be beneficial as it would allow us to create personalised medicine

Question 2

the whole genome is too large to sequence so it is cleaved into fragments , what are these called and how are they produced ?

Answer 2

restrictive fragments , enzymes called restrictive endonucleases

Question 3

how do you sequence the DNA form the restrictive fragments ?

Answer 3

Cutting the original DNA with other restriction enzymes with different sequence selectivity will give new fragments whose sequences partly overlap the fragments from the first restriction enzymes. Sequencing the restriction fragments and identifying the regions that overlap the fragments from different restriction enzyme cleavage allows the complete DNA sequence to be determined.

Question 4

what method does the sequencing machines use to determine the human genome ?

Answer 4

the sanger method

Question 5

whats needed for the sanger method ?

Answer 5

template single strand of DNA , primer , DNA polymerase , the 4 deoxynucleotidetriphosphates ( dATP ,dCTP,dTTP,dGTP and also modified di - deoxynucleotidetriphosphates called ddNTP’s

Question 6

what are the modified di -deoxynucleotide triphosphates ddNTP’s needed for ?

Answer 6

these terminate the strand elongation as they don’t have the OH group present on the C3’ so no phophsodiester bond can form

Question 7

what is the gel electrophoresis used for ?

Answer 7

to determine where the ddNTP is for each fragment as this allows the base sequence to be determined for the restrictive fragment

Question 8

how can the sanger method be accelerated ?

Answer 8

by using fluorescent labelling , the terminal base of each fragment is identified by its characteristic fluorescence . A different colour will be used ofr each ddNTP’s

Question 9

what does UV light generate from water ?

Answer 9

hydroxy radicals

Question 10

when dioxygen is present what does UV light cause ?

Answer 10

peroxy radicals to form

Question 11

what can the free radicals produced cause ?

Answer 11

they can react with the negatively charged backbone of DNA which could cause the chain to cleave. If this happens on one strand then it is easily repaired, but breaks on adjacent strands in close proximity lead to separation and eventual cell death. Also the radicals can react with the nucleotide bases and cause them to be oxidised ( lose electrons).

Question 12

how can the use of transcription factors lead to cancer ?

Answer 12

they bind between the amino acid and DNA bases of DNA which prevents protein binding as the bases have been modified

Question 13

what is a photochemical reaction and what can this cause ?

Answer 13

this is when UV light causes 2 opposite pyrimidines bases to join or dimerise this causes errors during DNA replication as mutations occur

Question 14

can the results of photochemical reactions be reversed and how ?

Answer 14

yes by the use of enzymes

Question 15

how can the bases be altered ?

Answer 15

by UV light

Question 16

what is an alkylating agent ?

Answer 16

this is a chemical that transfers an alkyl group ( CH3) to a nucleophile such as a nucleotide base of DNA.

Question 17

give an example of an alkylating agent

Answer 17

dimethyl sulphate ( strong electrophile) that reacts with the N-7 of purines as it is not involved in hydrogen bonding so it has lone pair of electrons that are easily alkylated.

Question 18

what do the nitrogen and sulphur mustards have attached to them and what does this occur ?

Answer 18

chlorine and nitrogen which are both highly electronegative and so they both become positively charged so there’s 2 electrophilic positions. This means that it can react twice with 2 separate DNA strands.

Question 19

how are mustards a treatment against cancer ?

Answer 19

covalent crosslinks can occur between DNA strands so replication cannot occur

Question 20

are some chemicals always alkylating agents ?

Answer 20

no some become active or are modified during excretion for example vinyl chloride

Question 21

describe N -nitrosoureas

Answer 21

these are a selective anticancer agent which is biologically inactive until activation. they are selective as they target only cancer cells. They are activated as a solid tumour doesn’t receive enough oxygen so the environment becomes acidic which activates the anticancer agent.

Question 22

what is a chain terminator ?

Answer 22

this is a modified nucleotide that prevents the nucleic acid chain from growing during replication

Question 23

What is acyclovir ( zovirac) used for?

Answer 23

used to treat infectious diseases caused by a DNA virus such as herpes.

Question 24

how is the acyclovir activated in vivio ?

Answer 24

the hydroxyl group is converted to a triphosphate

Question 25

what is the aclyclovir triphosphate look similar to ?

Answer 25

guanine base ,

Question 26

how does aclyclovir cause chain termination ?

Answer 26

the acylcovir triphosphate is taken by DNA polymerase to from an enzyme - substrate complex that has no OH on C3.

Question 27

what is AZT used to treat ?

Answer 27

reterovirus HIV that leads onto aids

Question 28

whats does the AZT resemble ?

Answer 28

deoxythymidine but the C3 has been replaced with an azido group ( N3)

Question 29

what is AZT changed into in vivo by enzymes ?

Answer 29

5 - triphosphate which is misrecognised to be deoxythymidine triphosphate by the viral DNA polymerase

Question 30

how does this affect chain elongation ?

Answer 30

the AZT - 5 - triphosphate does not contain the OH on C3 so therfore cannot elongate the chain