Biology- Unit 7

Question 1

Describe how PCR would be carried out

Answer 1

1) Mixture of DNA sample, primers, nucleotides and TAQ DNA polymerase
2) Mixture heated to 95 degrees to break hydrogen bonds between complimentary bases
3) Cooled to 50-65 degrees so primers can attach
4) Temperature increased to 70 degrees- TAQ Polymerase catalyses the polypeptide bonds between nucleotides
5) Cycle is repeated around 30 times or until one of the factors is used up

Question 2

Describe how DNA sequencing would be carried out

Answer 2

1) DNA sample divided into 4 separate sequencing reactions- all contain nucleotides, DNA polymerase, primers, and fluorescently tagged terminator nucleotides
2) DNA fragments of different lengths are formed across reaction vessels
3) High resolution gel electrophoresis is used to separate fragments by size
4) Fragments are visualised under UV, base sequence can be read

Question 3

Describe how DNA profiling would be carried out

Answer 3

1) Fragments of DNA are cut using restriction enzymes on either side of satellite regions
2) Fragments separate using gel electrophoresis, dyed with ethidium bromide so they fluoresce. Current is applied to gel, separate according to mass
3) Southern blot- alkane buffer solution added, nylon filter, fragments visible as blots, complimentary sequences labelled with fluoresce added
4) Blots compared to number of satellites visualise as graph, introns compared, more closely related = similar intron repeats

Question 4

What are the factors that affect gene expression?

Answer 4

Transcription factors
DNA methylation
Histone modification
Destruction of mRNA
mRNA splicing

Question 5

What dos DNA methylation entail?

Answer 5

A CH3 group is added to a Cytosine- prevents transcription factors binding

Question 6

What does Histone acylation entail?

Answer 6

Addition of an acetyl group which activated chromatin, ‘unwinds’ DNA from histone and allows DNA polymerase and transcription factors to bind

Question 7

What does mRNA splicing entail?

Answer 7

Pre-mRNA spliced
Introns removed
Exons can be rearranged in any order

Question 8

What is a multipotent stem cell?

Answer 8

Give rise to multiple types of cells

Question 9

What is a pluripotent stem cell?

Answer 9

Able to give rise to many types of specialised cells but not placental cells

Question 10

What is a totipotent stem cell?

Answer 10

Can give rise to all types of specialised cells

Question 11

How are induced pluripotent stem cells made?

Answer 11

1) Fibroblasts taken from skin samples
2) viruses used as vectors to introduce four genes for transcription factors
3) cells behave similarly to embryonic stem cells however can turn cancerous

Question 12

How is recombinant DNA produced?

Answer 12

Isolation of gene-
Section of DNA is cut out using restriction enzymes- leaves sticky ends
Use reverse transcriptase to make DNA from mRNA

Isolation of plasmid-
Cut with the same restriction enzymes used to leave complimentary sticky ends

Join plasmid and gene with DNA ligase to form recombinant DNA

Reincorporate plasmid into host nucleus

Question 13

What other vectors can be used to form recombinant DNA?

Answer 13

Gene guns
Viruses
Liposome wrapping
Microinjection

Question 14

How are transgenic plants made and what are their uses?

Answer 14

Genetic modification using agrobacterium tumefaciens

1) Ti plasmid extracted
2) Bacterial genes are inserted into plasmid via genetic modification
3) Plasmid is returned to bacterium
4) Plant is infected with bacterium
5) Plant grows a crown gall- contain inserted gene
6) Cells can be taken and cultured to grow whole plants

Uses-
Flood resistance
Pesticide production
Herbicide resistance
Changing nutrient value