PCR & Sequencing

polymerase chain reaction (PCR)

• produce many copies of a specific target segment of DNA

thermocycler

• dNTPs (free nucleotides)

• Taq polymerase

• buffer• 2 DNA primers

Figure 13.25

3

5

Cycle 1yields 2 molecules

Genomic DNA

Denaturation

Target sequence

3

5

3

5

3

5

Primers

New nucleotides

Annealing

Extension

Cycle 2yields 4 molecules

Cycle 3yields 8 molecules;

2 molecules(in white boxes)

match target sequence

Technique

1

2

3

PCR

http://www.dnalc.org/resources/animations/pcr.html

PCR song: ~2minhttp://www.cnpg.com/video/flatfiles/539/

DNA Sequencing

• Relatively short DNA fragments

• Can be sequenced by the Sanger dideoxy chain-termination method

http://www.dnalc.org/resources/animations/cycseq.html

Human Genome Project 1990-2003

Craig Venter’sshotgun-sequncing

-1000bp per day in 1980s-1000bp per second in 2000-100 billion bp per day in 2010!