Sequencing Genomes Dr Hilal AY DNA sequencing Chaintermination

Sequencing Genomes Dr. Hilal AY

DNA sequencing

Chain-termination Sequencing

Chain-termination Sequencing

Chain-termination Sequencing

Chain-termination Sequencing

Are all DNA polymerases suitable for sequencing? High processivity Negligible or zero 5ʹ → 3ʹ exonuclease activity Negligible or zero 3ʹ → 5ʹ exonuclease activity

Chain-termination sequencing with Taq polymerase Figure 4. 5 Different types of primers for chain-termination sequencing. (A) Forward, reverse, and internal primers enable different sections of a PCR product to be sequenced. (B) A universal primer anneals to the vector DNA adjacent to the position at which new DNA is inserted. A single universal primer can therefore be used to sequence any DNA insert.

Chain-termination sequencing

Strengths and limitations of chain-termination sequencing Figure 4. 6 read depth is needed in order to identify sequencing errors. Each region of a genome must be sequenced multiple times, in order to identify errors present in individual sequence reads. In this example, the discrepancy in read 4 in the highlighted column can be ascribed to a sequencing error, the correct nucleotide at this position being C.

NEXT-GENERATION SEQUENCING Roche 454 Illumina (Solexa) Ion torrent: Proton / PGM SOLi. D

NEXT-GENERATION SEQUENCING

Roche 454 pirodizileme

Roche 454 pirodizileme

Illumina/Solexa

Illumina/Solexa

Illumina/Solexa

Ion Torrent: Proton/PGM

Ion Torrent: Proton/PGM

Ion Torrent: Proton/PGM

ABI/SOLi. D

ABI/SOLi. D

Nanopore sequencing

Sanger sequencing– Next generation sequencing

Whole genome sequencing

Whole genome sequencing

Single cell genome sequencing

Single cell genome sequencing

Mitochondrial genome sequencing

Sequencing for genetic disorders

Sequencing for genetic disorders