Gene Technologies

Question 1

What is Recombinant DNA

Answer 1

• form of genetic engineering
• involves the transfer of fragments of
  DNA from one organism to another via isolating fragments of interest

Question 2

Why is genetic information transferable between species?

Answer 2

Genetic code is universal therefore the same amino acid code for the same amino acids in all living things

Question 3

What are the 3 methods to create fragments of DNA?

Answer 3

1. Reverse Transcriptase (enzyme)
2. Restriction endonucleases (enzyme)
3. Gene machine

Question 4

What is Reverse Transcriptase?

Answer 4

This process uses reverse transcriptase to make DNA copies of mRNA that was transcribed from the gene of interest

Question 5

What is the process of Reverse Transcriptase?

Answer 5

1. A cell that naturally produces protein of interest is selected
2. Once mRNA is isolated reverse transcriptase joins DNA nucleotide w/ complementary base pairs to the mRNA sequence
3. the mRNA is used as template to make the single stranded complementary DNA
4. DNA polymerase is then used to convert the single stand of cDNA into double stranded which contains the desired code for the gene

Question 6

Advantage of Reverse Transcriptase

Answer 6

• mRNA is present in cell is from transcribed genes → so lots of the mRNA of interest is available to make cDNA

Question 7

Disadvantage of Reverse Transcriptase

Answer 7

• more steps, so more time consuming & more difficult

Question 8

What is restriction endonucleases?

Answer 8

• this process uses the enzyme restriction endonucleases to cut up the DNA
• RE naturally occur in bacteria as a defence mechanism (if there is foreign DNA inside a bacteria, the enzyme will cut it up so it doesn’t replicate and cause harm)

Question 9

What is the process of restriction endonucleases?

Answer 9

1. RE will separate the 2 strands of DNA at a specific base sequence by cutting the sugar-phosphate backbone to give sticky ends or **blunt ends **
   BLUNT ENDS:
   - caused be being cut straight across
   - do NOT result in exposed bases
   STICKY ENDS
   - caused by cutting the DNA staggered
   - result in exposed DNA bases
   - palindromic (read the same front to back)
2. At sticky ends, the exposed base pairs can be aligned to exposed bases of the organism you want to insert it in → makes it easier to insert the desired gene into another organisms DNA
   can easily form H bonds w/ complementary base sequences on other pieces of DNA that have been cut w/ same restriction enzyme

Question 10

Advantage of Restriction Endonuclease

Answer 10

• Sticky ends on DNA fragment make it easier to insert in the DNA fragment when creating recombinant DNA
• can select enzymes that’ll cut at palindromic recognition sites to give sticky ends

Question 11

Disadvantage of Restriction Endonucleases

Answer 11

• Still contains introns as its cut from the original DNA–> an extra step is required to remove introns

Question 12

What is the gene machine?

Answer 12

using computers to generate the nucleotide sequence to produce the gene

Question 13

What is the process of the gene machine?

Answer 13

1. Scientists examine the protein of interest to identify the amino acid sequence
2. The DNA sequence is entered into the computer (this checks that he DNA being created is safe and ethical to produce)
3. The computer can create small sections of overlapping single strands of nucleotides that make up the gene (aka oligonucleotides)
4. Oligonucleotides can then be joined to create the DNA for the entire gene
5. Polymerase Chain Reaction can be used to make lots of copies of the gene and make it double stranded

Question 14

Advantages of Gene Machine

Answer 14

• very quick, very accurate
• can design exact DNA fragment you want with sticky ends, DNA probes & won’t have introns

Question 15

Disadvantages of Gene Machine

Answer 15

• need to know the sequence of amino acids or bases
• time consuming

Question 16

What is a DNA probe?

Answer 16

Short, single stranded prices of DNA with complementary base pairs.

Question 17

What makes DNA probes identifiable?

Answer 17

• Radioactively labelled probes
  -Fluorescently labelled probes

Question 18

?

Answer 18

1) Design DNA Probe w/ complementary sequence to allele being screened for
2) patients double stranded DNA is treated to separate and become single stranded
3) mis DNA with the probe binds can be identified
4)he ste at which the

Question 19

use of gene techinolgies

Answer 19

• allows screening for potential genetic disorders
• can identify presence of oncogenes
• can screen for multiple disease simultaneously using an array where multiple DNA probes are present

Question 19

How to locate alleles of genes?

Answer 19

1) must know DNA base sequence in order to design the probe
2) Could use DHNA sequencing techniques
3) produce DNA fragments 0 enzyme methos or gene machine
4)amplify- in vitro or in vivo cloning
5) add label- radioactive nucleotide or fluorescent label
6) presence of label indicates allele of intertest is present in patient DNA

After hybridisation , DNA is washed so any unbound probes are washed away

Question 19

why do we use gene technologies?

Answer 19

1. drug screening -some drugs are more/ less effective depending on genotype
2. cam help determine effective dose
   - saves money
   - increased effectiveness
   - increase safety
   -less waste