Genome Sequencing - Human Genome Project Flashcards
when did the human genome project begin?
October 1, 1990
how long did this project take?
13 years
who funded it?
U.S. Department of Energy and the National Institutes of health
what DNA was used?
human DNA that came from several volunteers
what were the goals of the human genome project?
1) obtain a genetic linkage map of the human genome
2) obtain a physical map of the human genome
3) obtain DNA sequence of the entire human genome
4) develop technology for the management of human genome information
5) analyze the genomes of model organisms
6) develop programs focused on understanding and addressing ethical, legal and social implications of the results obtained from the human genome project
7) develop technological advances in genetic methodologies
when was the first draft of the nearly complete sequence published? the second?
- February 2001
- 2003
when was the final draft published?
2006
how big is the human genome?
3 billion bp in length
high-throughput sequencing
ability to rapidly sequence large amounts of DNA
what are some advances that have made rapidly sequencing DNA possible?
1) fluorescently labeled nucleotides and detectors
2) parallel sequencing
next generation sequencing technologies
newer DNA sequencing tehcnologies that are more rapid and inexpensive than the dideoxy method
pyrosequencing
relies on the release of pyrophosphate, sis for the Roche 454/FLX
pyrosequencing steps
1) DNA is broken into small fragments of 300-800 bp
2) short oligonucleotides called adapters are linked to the 5’ and 3’ ends of DNA fragements
3) DNA is denatured to single strands
4) mixture of single strand DNA fragments with adaptors is called the sample library - single strands and beads attach via red adaptors (one strand per bead)
5) emulsify the beads so there is only one bead per droplet, droplets also contain pcr reagents that amplify the DNA
in pyrosequencing, what happens after droplets are formed?
each bead is placed into a well of a picoliter plate, where sequencing reagents are added to the wells
what does the picoliter plate do?
acts like a flow cell in which the beads are stuck in thew wells an pure solutions of different nucleotides can flow over them
- this is real time DNA