Lecture 2: Components Of Higher Eukaryotic Genomes Flashcards
What does the c-value paradox explain?
Genome size is not equal to genome complexity
What influences Cot value?
DNA concentration, translocation temperature, cation concentration, and viscosity
What is the Cot equation?
Co x t x buffer factor
Co = initial DNA concentration (mol/l)
t = renaturation time (sec)
Buffer factor = accounts for affect of cations on the speed of renaturation
What’s the purpose of Cot renaturation curves?
To determine genome complexity
What does a lower Cot value indicate?
DNA is more abundant and repetitive
What do protein coding genes contain?
~25kb consisting of introns and exons to be transcribed into RNA
What changes are made to protein coding gene to for mature mRNA?
- introns are spliced out
- no CpG island
- still contains exons and 5’ and 3’ untranslatable regions
How large is the genes translatable coding sequence and what does this tell us?
1.3kb, much less than 25kb
Shows that most of the unique parts of a gene are actually non coding
What are gene families?
Genes present in more than 1 copy per haploid genome which arise by duplication
What are the 3 different dates of duplicated genes?
Pseudogenization: gene inactivation
Neofunctionalization: evolution of a new function
Subfunctionalization: function is divided between genes
What is the difference between win structure between adult and foetal haemoglobin?
Adult subunits - 2 alpha and 2 beta
Foetal subunits - 2 alpha and 2 gamma
Where are haemoglobin subunits found?
Alpha subunits - tip of short arm of chromosome 16
Beta, gamma, and delta subunits - chromosome 11
Where is the largest histone gene cluster found in humans?
Chromosome 6
Why is it beneficial to have so many histone repeats?
Histones are important for DNA packaging, so lots of proteins are needed to carry out the function quickly
What are interspersed repeats?
Repeats dispersed throughout the genome, non adjacent to eachother
What are SINEs? How large are they and which is most abundant?
Short interspersed nuclear elements
100-300bp
Alu is most abundant, 11% of genome
What are LINEs? How large are they and which is most abundant?
Long interspersed repeats
1-5kb
Li is most abundant, 17% of genome
How do interspersed repeats use reverse transcription to spread?
Active LINEs encode reverse transcriptase, which then transposes themselves and SINEs to integrate
What causes the intron 22 inversion mutation of the F8 gene?
3 LINE insertions
What are VNTRs?
Variable number tandem repeats which are dispersed throughout the genome and are short sequences repeated multiple times in tandem
What are the 3 groups of VNTRs and their sizes?
Satellite DNA: >100bp located in centromere (AT rich)
Minisatellite DNA: 15-100bp
Microsatellite DNA: <15bp
Why is the important micrsatellite DNA found in telomeres?
Hexameric sequence TTAGGG
Why are VNTRs variable in number?
- unequal crossing over
- DNA replication slippage
- DNA repair
- repeat number is inherited through pedigrees
What causes triplet repeat diseases?
Length variation in specific microsatellites, as expansion beyond a threshold results in morphological disease
What are the steps involved in DNA fingerprinting?
- genomic DNA is extracted from biological sample
- DNA is digested into small fragments by restriction enzymes
- some fragments contain mini satellites of interest
- electrophoresis separates fragments by size
- transferred to membrane using southern blot
- membrane is incubated with fluorescent DNA probes and hybridize with complementary fragments
- results show pattern of bands on gel
What do probes of minisatellite sequences detect?
Polymorphic patterns of restriction endonuclease fragments
How can genetic fingerprints be generated using multiple microsatellite VNTRs
- specific parts of DNA are put though PCR with PCR primers
- amplifies different microsatellites which are labelled with coloured fluorophores
- labelled DNA fragments are generated and separated with electrophoresis
What is genetic fingerprinting used for?
Paternity tests, forensics, identifying diseases and wildlife conservation studies
