Progress in genome sequencing Human Genome Project 10

Progress in genome sequencing · Human Genome Project · 10 years to complete · Billions of dollars · Current sequencing technology · $10 -25 million to sequence a human genome · Mammalian genomes sequenced in months · Microbial genomes sequenced in weeks · Massively parallel sequencing · 25 million base pairs in 4 hours! © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

Sample preparation · Fragment DNA into short single strands · Use “adaptor” sequences to attach DNA to micron -scale beads · Encase beads in oil droplets containing PCR reagents · Amplify bead DNA · Load beads into picoliter wells From Figure 1 in Margulies, M. et al. (2005) “Genome sequencing in microfabricated high-density picolitre reactors” Nature 437: 376 -380. Figures 1 and 2 used by permission by Jonathan Rothberg. © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

The instrument · Load beads into slide containing 1. 6 million picoliter wells · Sequentially pass A, G, C, and T into picoliter wells · Presence of a particular base emits light from individual well · CCD reads emitted light from each picoliter well From Figure 2 in Margulies, M. et al. (2005) “Genome sequencing in microfabricated high-density picolitre reactors” Nature 437: 376 -380. © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

Caveats · Only reads short lengths of sequence · 80 -120 bp · Accuracy of individual reads is lower than conventional methods · Though speed can compensate to some extent · Can not read paired-ends · Makes sequence assembly less efficient © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458