Gene Technology- Genome Projects and Making DNA Fragments

Question 1

What is a genome?

Answer 1

The entire set of DNA, including all the genes in an organism

Question 2

What have improvements in technology allowed us to do?

Answer 2

To sequence the genomes of a variety of organisms

Question 3

What do gene sequencing methods only work on?

Answer 3

Fragments of DNA which are sequenced and then put back in order to give the sequence of the whole genome

Question 4

What was the Human Genome Project and when was it completed?

Answer 4

Mapped the entire sequence of the human genome for the first time in 2003

Question 5

What is a proteome?

Answer 5

All the proteins that are made by an organism

Question 6

What type of organisms don’t have much non-coding DNA?

Answer 6

Simple organisms such as bacteria

Question 7

Why is it helpful when organisms don’t have much non-coding DNA?

Answer 7

It is relatively easy to determine their proteome from the DNA sequence of their genome

Question 8

When can it be useful for organisms not having much non-coding DNA?

Answer 8

In medical research and development

Question 9

Why is it harder to translate the genome of complex organisms?

Answer 9

They contain large sections of non-coding DNA and they also contain complex regulatory genes (determine when the genes that code for particular proteins should be switched on or off)

Question 10

Why does containing large sections on non-coding DNA make it more difficult to translate the genome of complex organisms into their proteome?

Answer 10

Because it’s hard to find the bits that code for proteins among the non-coding and regulatory DNA

Question 11

What were sequencing methods like in the past?

Answer 11

Labour-intensive, expensive and could only be done on a small scale

Question 12

What are sequencing methods like now?

Answer 12

Automated, cost-effective and can be done on a large scale

Question 13

What does recombinant DNA technology involve?

Answer 13

Transferring a fragment of DNA from one organism to another

Question 14

What are organisms that contain transferred DNA known as?

Answer 14

Transgenic organisms

Question 15

What are the three ways that DNA fragments can be produced?

Answer 15

Using reverse transcriptase, using restriction endonuclease enzymes and by using a gene machine

Question 16

Why is it difficult to obtain a DNA fragment containing the target gene?

Answer 16

Most cells only contain two copies of each gene

Question 17

Why is mRNA easier to obtain?

Answer 17

Cells contain many mRNA molecules which are complementary to the gene

Question 18

How can mRNA molecules be used?

Answer 18

As templates to make lots of DNA

Question 19

What does reverse transcriptase do?

Answer 19

Makes DNA from an RNA template

Question 20

What is the DNA called that is produced from an RNA template?

Answer 20

cDNA- complementary DNA

Question 21

What do pancreatic cells have?

Answer 21

Loads of mRNA molecules complementary to the insulin gene, but only two copies of the gene itself

Question 22

What can reverse transcriptase be used for in pancreatic cells?

Answer 22

Making cDNA from the insulin mRNA

Question 23

How does reverse transcriptase make cDNA from the insulin mRNA?

Answer 23

• mRNA is isolated from cells
• Mixed with free DNA nucleotides and reverse transcriptase
• The reverse transcriptase uses the mRNA as a template to synthesise a new strand of cDNA

Question 24

What are palindromic sequences of nucleotides?

Answer 24

Sequences that consist of antiparallel base pairs

Question 25

What are restriction endonucleases?

Answer 25

Enzymes that recognise specific palindromic sequences and cut the DNA at these places

Question 26

Why do different restriction endonucleases cut at different specific recognition sequences?

Answer 26

Because the shape of the recognition sequence is complementary to the enzyme's active site

Question 27

When can you use restriction endonucleases to separate the fragment from the rest of the DNA?

Answer 27

If recognition sequences are present at either side of the DNA fragment you want

Question 28

How is the restriction endonuclease used?

Answer 28

The DNA sample is incubated with the specific restriction endonuclease which cuts the DNA fragment out via a hydrolysis reaction

Question 29

What can restriction endonucleases leave?

Answer 29

Sticky ends

Question 30

What are sticky ends?

Answer 30

Small tails of unpaired bases at each end of the fragment

Question 31

What can sticky ends be used for?

Answer 31

To bind (anneal) the DNA fragment to another piece of DNA that has sticky ends with complementary sequences

Question 32

How has technology recently been developed?

Answer 32

So that fragments of DNA can be synthesised from scratch without the need for a pre-existing DNA template

Question 33

What does the database involved in gene machine contain?

Answer 33

The necessary information to produce the DNA fragment

Question 34

How does the gene machine work?

Answer 34

- Sequence required is designed if it doesn't already exist - First nucleotide in the sequence is fixed to some sort of support - Nucleotides are added step by step in the correct order, in a cycle of processes the includes adding protecting groups - Oligonucleotides are produced - Broken off from the support and all the protecting groups are removed - Oligonucleotides can the be joined together to make longer DNA fragments

Question 35

What do protecting groups do?

Answer 35

Make sure that nucleotides are joined at the right points, to prevent unwanted branching

Question 36

What are oligonucleotides?

Answer 36

Short sections of DNA