Videos- Recombinant DNA Tech

Question 1

3 ways to isolate target genes

Answer 1

1) Restriction enzymes
2) Reverse transcriptase
3) Artificial synthesis of gene

Question 2

What does palindromic mean?

Answer 2

Sequence reads same forward and backwards

Question 3

What does reverse transcriptase do?

Answer 3

Enzyme that does transcription backwards- mRNA➡️ cDNA

Question 4

What’s cDNA?

Answer 4

Complementary DNA that has no introns made using reverse transcriptase

Question 5

What’s a digonucleotide?

Answer 5

Nucleotides joined together during the process is artificial synthesis of a gene (max 25)

Question 6

What’s the process of making a gene from scratch? (Artificial synthesis gene)

Answer 6

1) Use gene machine.

2) Form digonucleotide, join multiple digonucleotides together to make a synthetic gene.

Question 7

How to isolate a target gene using restriction enzymes?

Answer 7

Restriction enzymes cut DNA at restriction sites. Leaves DNA with sticky ends.

Question 8

What’s a restriction site?

Answer 8

Specific palindromic site

Question 9

3 main steps in forming a transgenic bacteria

Answer 9

1) Isolate target gene
2) Inset gene into vector
3) Insert vector into bacteria

Question 10

4 things needed for isolating target genes

Answer 10

1) Promoter region
2) Terminator gene
3) Sticky ends
4) Marker genes

Question 11

What’s a vector?

Answer 11

Something used to move DNA from one place to another

Question 12

2 types of vectors

Answer 12

Virus- bacteriophage

Plasmid- double stranded loop of DNA, transfers genes between bacteria

Question 13

How do you insert a gene into a vector?

Answer 13

Use restriction enzyme to cut plasmid =complementary sticky ends
DNA ligase reforms phosphodiester bonds
Forms

Question 14

How do you insert a gene into a vector?

Answer 14

Use restriction enzyme to cut plasmid =complementary sticky ends
DNA ligase reforms phosphodiester bonds
Forms recombinant DNA

Question 15

How do you insert the vector into the bacterium?

Answer 15

Ice cold CaCl2 and heat shock it

Question 16

What’s formed when a vector is inserted into a bacterium?

Answer 16

Transgenic organism

Question 17

Why are marker genes used in recombinant DNA?

Answer 17

To identify the transformed bacteria

Question 18

What’s a marker gene? (According to tailored tutors)

Answer 18

Easy to identify

Paired with target genes to see whether the vector has been incited properly

Question 19

2 examples of marker genes

Answer 19

1) UV fluorescence

2) Antibiotic resistance

Question 20

What does the polymerase chain reaction do?

Answer 20

In vitro amplifies DNA, makes lots of copies

Question 21

What do you need for PCR? (polymerase chain reaction)

Answer 21

DNA sample
Free DNA nucleotides
Primer
DNA polymerase

Question 22

What’s the method of PCR?

Answer 22

1) Heat to around 95 degreesC
2) Cool to around 50 degreesC
3) Heat to around 70 degreesC
4) Repeat.

Question 23

What’s the reason for the first step in PCR? (Heating to around 95 degreesC)

Answer 23

Break hydrogen bonds

Make DNA single stranded

Question 24

What’s the reason for the second step in PCR? (Cool to around 50 degreesC)

Answer 24

Allows primer to bind
Complementary base pairing
DNA double stranded
DNA polymerase can bind

Question 25

What's the reason for step 3 in PCR? (Heat to around 70 degreesC)

Answer 25

DNA polymerase adds complementary nucleotides | Makes phosphodiester bonds

Question 26

What's the reason for repeating the process of PCR? (4th step)

Answer 26

To make as much DNA as possible, each cycle doubles the amount of DNA

Question 27

What is electrophoresis?

Answer 27

Using electricity to seperate DNA fragments by length

Question 28

What's the process for electrophoresis?

Answer 28

Attach fluorescent label/stain DNA fragment Put marked DNA fragments in well at anion Turn on current DNA is negative so is attracted to cation Smaller fragments move faster so further as there's less resistance.

Question 29

How do you calibrate your scale for electrophoresis?

Answer 29

Using known lengths of DNA

Question 30

Summary of gene technology process

Answer 30

1) isolate target gene 2) insert gene into vector 3) insert vector into bacterium 4) identify transgenic bacteria 5) culture transgenic bacteria 6) extract and purify protein

Question 31

Who do you culture your transgenic bacteria?

Answer 31

Transcribe and translate recombinant DNA | Make proteins of target gene

Question 32

What's the purpose of recombinant DNA and gene technology?

Answer 32

To make a protein from your target gene | E.g. Insulin

Question 33

What is gene therapy?

Answer 33

Changing faulty alleles that cause genetic disease

Question 34

How do you change a faulty allele if the allele at fault is dominant? (Process)

Answer 34

Silence dominant allele 1) use vector to add DNA fragment into dominant allele 2) dominant allele isn't transcribed 3) by default recessive allele is expressed, (transcribed and translate)

Question 35

How do you change a faulty allele if the allele at fault is recessive? (Process)

Answer 35

Sufferer is homozygous 1) use vector to add functional allele to DNA- has to be added in right locus 2) dominant allele expressed

Question 36

What is germ line gene therapy?

Answer 36

Changing alleles of gametes

Question 37

What so somatic gene therapy?

Answer 37

Changing alleles of body cells

Question 38

How do somatic and germ line gene therapy compare with how they affect future offspring?

Answer 38

Germ line- offspring inherit changes | Somatic- offspring don't inherit changes

Question 39

What are 4 possible problems of gene therapy?

Answer 39

1) alleles inserted into wrong locus 2) silence wrong gene e.g. Tumour suppressor gene which would cause cancer 3) gene over expressed 4) used for non-medicinal reasons e.g. Designer babies or makeup

Question 40

What are the uses of genetic modification in agri- e.g. Soya beans?

Answer 40

Express protein from bacteria Protein toxic to insects Fewer insects on plants

Question 41

What are the advantages of genetically modifying soya beans?

Answer 41

Use less chemical pesticides | More effect food chain

Question 42

How and why is golden rice genetically modified?

Answer 42

Gene from corn p,ant inserted into rice plant | Allows VitA expression

Question 43

What's the advantage of genetically modifying golden rice?

Answer 43

Prevents blindness from VitA deficiency

Question 44

What are 3 disadvantages of genetically modifying plants in agriculture?

Answer 44

1) monoculture of crop with low genetic diversity- susceptible to disease/enviroment 2) buy new seed every year- termination gene 3) decreased biodiversity

Question 45

What does industry and research use genetic modification for?

Answer 45

1) making enzymes E.g. Re in (cheese) and lipase 2) transform pathogens to treat disease- attack other pathogens without infecting humans

Question 46

What's the advantage of genetically modified enzymes?

Answer 46

Reduced energy and cost | Fast and cheap production

Question 47

What's the advantage of genetically modified pathogens?

Answer 47

Treats disease Pathogens won't develop resistance Reduces suffering

Question 48

What are the disadvantages of genetically modified pathogens?

Answer 48

Could mutate and infect humans | Could be used in war as bio weapons

Question 49

What does medicine 'pharming' use genetic modification for?

Answer 49

Transform bacteria to express proteins | Transform mammals to produce useful products in their milk

Question 50

What's the advantage of genetic modification in medicine?

Answer 50

Make human proteins | Cheaper and easier than making synthetic proteins

Question 51

What are the disadvantages of genetically modified medicine?

Answer 51

Possible unexpected problems e.g. Cancer in mammals | Using animals as a commodity