BME 130 – Genomes
Lecture 4
Sequencing technology IINext generation
sequencing
Figure 4.2 Genomes 3 (© Garland Science 2007)
Figure 4.3a Genomes 3 (© Garland Science 2007)
454 Process Overview
1) Prepare Adapter Ligated ssDNA Library
2) Clonal Amplification on 28 µ beads
4) Perform Sequencing by synthesison the 454 Instrument of ~250,000 molecules
3) Load beads and enzymes in PicoTiter Plate™
Key sequence = TCAG for identifying wells and calibration
TACG
Flow Order
454 base calling
Limitations of Roche/454
Limitations of Solexa/Illumina
Limitations of ABI/SOLiD
Applications ofhigh-throughput sequencing
CHiP-Seq
Johnson et al., Science 8 June 2007:Vol. 316. no. 5830, pp. 1497 - 1502
RNASeq
Cloonan et al., Genome Biology 2008, 9:234