control of gene expression

Question 1

what is a gene mutation defined as

Answer 1

any change to one or more nucleotide bases or rearrangement of bases

Question 2

what can rate of cell division depend upon

Answer 2

environment, but also genes

Question 3

what could happen to cell division if gene mutation occurs

Answer 3

the rate of cell division is controlled by genes, if a mutation occurs in those genes then uncontrolled cell divison can occur. CANCER . when these cells keep divided they layer on top of each other forming a tumour

Question 4

what are proto-oncogenes

Answer 4

genes that can cause cancer when they are turned on

Question 5

what are tumour suppressor c

Question 6

what are three methods to produce DNA fragments

Answer 6

conversion of mRNA to cDNA using reverse transcriptase
- using restriction endonucleases to cut fragments containing the desire gene from DNA
- creating the gene in a gene machine based on a known protein structure

Question 7

describe the method to produce DNA fragments using reverse transcriptase

Answer 7

• the beta cells in the islets of langerhan are specialised to make insulin they therefore make a lot of mrna that codes for insulin
• this mrna will act as a template on which a single stranded complementary copy of DNA is made cDNA is formewd using reverse transcriptase
• cDNA is then isolated via hydrolysis of the mRNA using an enzyme
• double stranded dna is formed on the template of the cDNA using DNA polyermase
  this is a copy of the gene

Question 8

describe the mechanism of using a restriction endonucleases

Answer 8

• restriction endonucleases cute up DNA
• these naturally occur in bacteria as defence mechanisms
• each enzyme recognises a specific DNA sequence ( recognition site) which it is complementary to
• the DNA is cut by breaking phosphodiester bonds at the recognition site
• this will leave sticky ends whcih can easily add recombinant DNA

Question 9

how does the gene machine work

Answer 9

the primary structure of the DNA is examined to identify the amino acid sequence then mRNA and dna can be worked out

• the dna sequence can be entered in a computer which checks for biosafety and biosecurity
• the computer can then create small section of overlapping single stranded nucleotides called oligonucleotides
  the oligonucleotides can then be joined to create DNA for the entire gene
• using sticky ends the gene can then be inserted into a plasmid which acts as a vector for the gene allowing it to be stored, cloned or transferred in the future

Question 10

what is an oligonucleotide

Answer 10

small section of overlapping singled stranded nucleotides

Question 11

what is an advantage and disadnavntage of the reverse transcription metjod of making dna fragment

Answer 11

MANY NAYURALLY OCCURING MRNA ##

but this process is more difficult

Question 12

what are the adv and dadv of using endonucleases to form dna fragments

Answer 12

produces sticky ends so recombinant DNA can attach easily

produces introns - bacteria should not have introns

Question 13

what are advantages and disadvantages of gene machine

Answer 13

specific so that introns can be removed, quick

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Question 14

what is a palindrome

Answer 14

where two sequenecs are opposites of one another

Question 15

what does in vivo mean

Answer 15

tranferring the fragments to a host cell using a vector

Question 16

what does in vitro mean

Answer 16

uses the polymerase chain reaction

Question 17

what is a recognition site

Answer 17

the sequences of DNA that are cut by restriction endonucleases

Question 18

what does DNA ligase do

Answer 18

joins complementary sticky ends

Question 19

REMEMBER

Answer 19

USE THE SAME RESTRICTION ENDONUCLEASE TO BE ABLE TO COMBINE COMPLEMENTARY STICKY ENDS TOGTHER at plasmid and at dna

Question 20

what does the preparation of a DNA fragment involve

Answer 20

addition of extra lengths of DNA

Question 21

what is a promoter region

Answer 21

the binding site of RNA polymerase whcih will allow transcription to occur

Question 22

what is terminator region

Answer 22

where another region of DNA releases RNA polymerase and stops transcription of the first little strand

obvs there will need to be another terminator region at the end of DNA to stop it all

Question 23

briefly describe gene transfer

Answer 23

• isolation of DNA fragments that have a gene for a desired protein
• insertion of DNA fragment into a vector
• transfer the DNA into a suitable host cell
• identify host cells that ahve successfully taken up the gene by use of gene markers
• growth by cloning of the population of host cells

Question 24

what can be used as gene markers

Answer 24

fluorescence’s causes cells to glow under UV light
- bacterial cells which have not taken up the plasmid will not be antibiotic resistant

Question 25

what is a vector

Answer 25

a carrying unit

Question 26

what are vectors used for

Answer 26

used to transport DNA into the host cell

Question 27

where do restriction endonucleases usually break in the plasmid

Answer 27

the gene for antibiotic resistance

Question 28

describe the transformation process

Answer 28

plasmids and bacterial cells are mixed together in a medium containing calcium ions. the calcium ions and changes more permeable allowing the plasmids to pass through the cell surface membrane -- however not all the bacteria will posses the DNA of the desired gene

Question 29

what are some reason that the desired gene may not be taken up by bacterial cells

Answer 29

-some plasmids may have closed without taking in the DNA fragment - sometimes the DNA fragments join together to form its own plasmid

Question 30

what can PCR be described as

Answer 30

in vitro method of amplification

Question 31

what is the purpose of PCR

Answer 31

to produce large quantities of of DNA or RNA fragments

Question 32

what equipment is needed for PCR

Answer 32

- thermocycler - DNA fragment that intends to be amplified - DNA polymerase ( taq polymerase) - primers DNA nucleotides

Question 33

describe the process of PCR

Answer 33

- PCR mixture is heated to high temperature 95 degrees. this will break hydrogen bonds to separate the DNA strands - reduce the temperature ( 55 degrees ) so primers can anneal to DNA - heat to higher temp (72 degrees ) which is optimum temp for DNA polymerase - An enzyme adds adjacent nucleotides that will form phosphodiester bonds. this will make 2 stranded DNA repeat this cycle 25-25 times

Question 34

how many times should the PCR cycle be repeated

Answer 34

25-35

Question 35

what does heating the PCR mixture to 95 degrees do

Answer 35

breaks hydrogen bonds between DNA strand NO HYDROLYSIS

Question 36

what is a primer

Answer 36

a short single stranded DNA sequence

Question 37

what is the purpose of the PCR mixture being cooled

Answer 37

so primers can anneal to complementary bases at the end of DNA fragemnts

Question 38

what is the purpose of PCR

Answer 38

DNA amplification

Question 39

describe genetic fingerprinting

Answer 39

DNA is extracted by fractionation and ultracentrifugation ( if the sample is super small the PCR is used to amplify DNA sample) - restriction endonucleases are added to cut the DNA into fragments - these samples are added to wells in agar gel. - the gel is placed in a buffer liquid with an electrical voltage applied - DNA is negatively charged therefore the DNA samples move through the gel towards the positive end of the gel - -the agar gel creates resistance for the moving DNA, and smaller pieces of DNA can move faster and further along the gel - this is how different lengths of DNA are separated (VNTRs) ahhhhhhhhh that bit was gel electrophoresis an alkaline is added to separate the double strands of DNA

Question 40

Why is an alkaline added after gel electrophoresis

Answer 40

to separate the double strands of DNA

Question 41

what can genetic fingerprinting be used for

Answer 41

to analyse genetic relationships and genetic variability within a population

Question 42

what is a VNTR

Answer 42

variable number tandem repeats - found in the non-coding region of DNA. between individuals contain variable lengths of repeating DNA

Question 43

how do you analyse genetic fingerprinting

Answer 43

add a DNA probe which is a single stranded piece of DNA complementary to VNTRs - these probes are radioactively or fluorcescently labelled - the probes are mixed in with the agar gel - they hybridise - the agr gel will shrink and crack as it dries. the VNTRs and DNA probes are transferred to a nylon sheet. this nylon sheet is exposed to X-rays to visualise the position of the radioactive probes or UV is a fluorescent probe was used DNA bands are then compared to identify geentic relationships