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Personal Genome Project (PGP) Goal: sequencing full genome of individuals at $1000
Human Genome Project (HGP) (total 3 billion $) : Motivated 100X reduction in cost (10 $ per base to 10 base per 1$.) Personal Genome Project(PGP: has motivated the development of Ultra-Low-Cost-Sequencing (ULCS).
Pyrosequencing
http://mpss.udel.edu/tutorial/flash/megaclone.swf
Megaclone transforms a sample containing millions of DNA molecules into one made up of millions of micro-beads, each of which carries approximately 100,000 copies of one of the DNA molecules in the sample.
Nanopore sequencing
Single stranded polynucleotides can only pass single file through a hemolysin nanopore. The presence of the polynucleotide in the nanopore is detected as a transient blockade of the baseline ionic current. PA-pico-Ampere
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Website:
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Fig. 11.3
Microsatellites
1 every 30,000 bp Repeated units 2 5 bp in length Mutate by replication error Mutation rate of 10-3 per locus per gamete Useful as highly polymorphic DNA markers
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Minisatellites
Repeating units 20-100 bp long Total length of 0.5 20 kb 1 per 100,000 bp, or about 30,000 in whole genome
Fig. 11.4
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Ler Col
Probe:
HindIII digestion Electrophoresis Southern blotting Hybridization with the probe autoradiography
1kb ladder Ler
Col Het
2 1
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Fig. 11.6
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Example of RFLP
Restriction digestion GE Ethedium bromide stain Blotting Hybridization X-ray radiography
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Ler
HindIII
2 kb
HindIII
5.7 kb
HindIII
HindIII
Col
LCM
5
2 1
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Fig. 11.7
CAPS
Must have sequence on either side of polymorphism Amplify fragment Expose to restriction enzyme Gel electrophoresis e.g., sickle-cell genotyping with a PCR based protocol
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