Human Genome Project Flashcards
Human Genome Project
Was a commitee that sought to sequence the human genome, and eventually of other organism
Who did the HGP?
1988 by the US national academy
Original issues with the HGP?
Ethical issues, low computing power and vast amoutn of genes where sanger sequencing was limited.
What were the two groups working on the HGP?
International HGP(HGS approach)
Celera Genomes(WGS approach)
Hierarchical Genome Sequencin
Analysis entireity of the human genome in a cell at a single time.
What did HGS use?
RE to cleave human genome ito 200,000 fragments and BAC
Process of HGS
Break genome into smaller fragments(100,000NT pairs)
Ordered clone maps(comparison of restriction enzyme cleavage sites in a given BAC clone
Using GE to sequence ordered clonse, generating whole genom using PCR
COmputs alligning sequence to figure out where the chromsomes go
What does WGS sequencing use instead?
Directly taking DNA into short fragments, ligated into plasmids
Process of WGS
Ligating into fragments, each fragment sequenced, ordered in a computer softwar
Why was WGS slower?
Computing power required due to not physically ordering the framgnets
Differences of WGS and HGS
Hgs physically ordered whislt WGS did not
HGP by 2003?
90% of human genom sequence
Why was 100% sequencing hard?
Lots of repetetive sequences hard to allign together(especially with sanger sequencing)
When was HGP completed?
2022 due to 2nd and 3rd generation sequencing
Why were newer techniques better in sequencing?
Not requiring DNA libraries and use of PCR instead of BAC
Illumnia Sequencing
This prepares DNA libraries by binding DNA to solid surface then amplification by PCR
What is sequencing done with?
dNTP and fluorophores, so each Polymerase NT addition the machine reads light emitted, detemrining order of each dNTP
Light emission depending on NT joined…
If NT joined to growing strand, PPI is released causing a flash of light, whilst if not complemnetary, no reaciton thus no flash.
Ion Torrent Sequncing
Uses welds containing beads coated each with DNA, each with a different sequence
Differnce between Ion Torrent Sequencing and Illumnia Sequencing?
Everytime dNTP is added, machine reads change in PH instead of fluorophores, as each dNTP added releases a proton
Types of 2nd generation sequencing…
Illumnia and Ion Torrent Sequencing
Examples of 3rd generation sequencing?
PACbio and Oxford Nanopores
What can a genome sequence be used for?
Analysis of gene mutaiton for cancer tpyes
Analysis of parasitic pathways using known sequences of genes in humans
Identify genes in human genome compare with other species for eovlutonary genetics
How are protein coding genes ID?
A protein has a reading frame with start/stop codon
Promoter sequences
RNA-splicing sites(introns/exons)
The point of the 1000 genome projcct in 2008?
Between two humans, 99% of genome is identical, so studying variation to identify susceptibility to diseases
Where HAS genome sequencing been used?
Cancer genomics, identification of mutations, in agriculture for crop seqeuncing, forensics, microbiomes and microbes
