Human Genome

Question 1

How much of the genome encodes for proteins ?

Answer 1

5%

Question 2

What are Isochores ?

Answer 2

Large DNA segments (>300 kb) which are characterised by an internal variation in GC content

Question 3

Describe CpG Islands

Answer 3

• Cytosine base followed by a guanine base is rare In vetebrate DNA
• Cytosines following guanines tend to be methylated - the methylation state of these CpG islands can regulates the expression of the genes

Question 4

Describe LINE retroansposon

Answer 4

• repetitive element in genome
• retro = going through an RNA intermediate
• LINE = Long Interspersed Elements
• complete sequence is 6000-8000 bp long

Question 5

Describe a SINE retrotransposon

Answer 5

• SINE = Short Interspersed Elements
• its a parasite’s parasite –> depends on LINE for its propagation
• Alu elements are most abundant in humans - 300bp long

Question 6

How much of the genome are interspersed repeats ?

Answer 6

46%

Question 6

What are the 4 repetitive elements found in the human genome ?

Answer 6

1. LINE retrotransposon
2. SINE retrotransposon
3. Retroviruses
4. DNA transposon

Question 7

What are Alu Elements ?

Answer 7

• only found in primates
• can be sorted into distinct families according to shared patterns of variation
• only one or several Alu “master copies” are capable of transposing

Question 8

What are flanking regions ?

Answer 8

• consist of ‘unordered’ DNA
• occur on each side of the repeat unit
• critical because they allow for the development of locus-specific primers to amplify the microsatellites with PCR

Question 9

Why are repetitive elements bad?

Answer 9

• repetitive elements waste energy
• insertion of of these repeats can be harmful

Question 10

Why do repetitive elements still exist ?

Answer 10

• generation & deletion of repeats have reached an equilibrium
• mammalian genomes can tolerate them as they’ve developed mechanisms to control them - eg histone modification

Question 11

Describe a duplicated pseudogene

Answer 11

• created from tandem duplication or unequal-crossover
• segment duplication is prevalent

Question 12

What is a multigene family ?

Answer 12

groups of genes from the same organism that encode proteins with a similar sequence either over their full length or limited to a specific domain

Question 13

What results in a multigene family ?

Answer 13

• DNA duplications that involve 1 or more genes generate gene pairs
• if both copies are maintained in subsequent generations then a multigene family will exist in the genome

Question 14

What are histone proteins?

Answer 14

• globular in shape
• 5 members in the family = H1, H2A. H2B, H3 and H4
• the family members are closely related but not identical in amino acid sequences

Question 15

Describe the structure of histone proteins

Answer 15

• octamer 2x H2A, H2B, H3 & H4
• H1 holds the structure together
• the globular protein contain tails which are rich in amino acids which have positively charged R-groups

Question 16

Describe epigenetics

Answer 16

• phenotype changes in a cell/organism
• not the result of nucleotide changes
• brought about by chemical changes to DNA (CpG) methylation and histones

Question 17

What is the first distinct mechanism caused by histone modifications and how does it affect chromosome function ?

Answer 17

modifications may alter the electrostatic charge of the histone resulting in a structural change in histones or their binding to DNA

Question 18

What is the second distinct mechanism caused by histone modifications and how does it affect chromosome function ?

Answer 18

modifications are binding sites for protein recognition modules, such as chrodomains that recognise acetylated lysines or methylated lysine

Question 19

What are 2 problems in genome sequencing ?

Answer 19

1. genomes are incredibly large
2. you need a lot of DNA to get a sequence

Question 20

What are the solutions to the problems of genome sequencing ?

Answer 20

1. cut the DNA into smaller pieces then put it back together
2. make lots of copies of each bit

Question 21

Describe PCR

Answer 21

• Polymerase Chain Reaction
• enables large amounts of DNA to be produced from very small/very complex samples
• uses 2 primers & DNA polymerase

Question 22

What does PCR allow?

Answer 22

• selecting the region of study
• producing millions of copies of it

Question 23

Describe Sanger sequencing

Answer 23

• similar to PCR
• uses only a single primer and polymerase to make new single stranded DNA pieces

Question 24

What is used to reduce the size of DNA in order to sequence it ?

Answer 24

• restriction enzymes
• they recognise and cut specific sequences
• there are hundreds of restriction enzymes are available

Question 25

What is BAC ?

Answer 25

• Bacterial Artificial Chromosomes
• after the use of restriction enzymes there is a soup of DNA fragments
• we use the BAC to amplify and identify each fragment

Question 26

What are the 2 purposes of the BAC libraries?

Answer 26

1. separates fragments so they can be sequenced individually
2. allows the production of lots of DNA by growing up lots of bacteria containing identical BACs or plasmids

Question 27

Why is BAC library not suitable for genome sequencing on a large scale ?

Answer 27

• 20,000 different BAC clones are needed to contain the 3 billion pairs of bases in the genome
• each inserts 150,000-200,000 base pairs
• minimum of 1.5 million sub-clones needed

Question 28

what are the simplified steps of DNA sample preparation ?

Answer 28

1. Extract DNA
2. Randomly Shatter (sonifcation)
3. Attach adapter sequence

Question 29

Describe Illumina Next-Generation Sequencing

Answer 29

• most popular method
• produces millions of reads
• reads can be paired-end
• very good for alignment

Question 30

Describe DNA Barcoding

Answer 30

• barcoded = tagged with a short known sequence
• allows multiple samples to be put on the same run
• samples can be computationally isolated