PCRPCRPolymerase Chain Polymerase Chain

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What is it and how is it What is it and how is it useful?useful?

A quick, easy method for amplifying (creating billions A quick, easy method for amplifying (creating billions of copies) of unimaginably tiny amounts of DNAof copies) of unimaginably tiny amounts of DNA

Methodology closely related to process of DNA Methodology closely related to process of DNA replicationreplication

Used in forensic science to analyze small quantities of Used in forensic science to analyze small quantities of DNA recovered from a crime scene such as:DNA recovered from a crime scene such as: - saliva on a cigarette butt or envelope- saliva on a cigarette butt or envelope- DNA in the root of a single strand of hair - DNA in the root of a single strand of hair - tissue under the fingernails of a victim - tissue under the fingernails of a victim - DNA in a single drop of blood- DNA in a single drop of blood- a small semen stain- a small semen stain

Used to detect disease/infection in an Used to detect disease/infection in an organism and treat it before a condition organism and treat it before a condition worsensworsens

Anthropology/archeology – PCR has Anthropology/archeology – PCR has allowed tiny fragments of DNA that allowed tiny fragments of DNA that have been exposed and degraded over have been exposed and degraded over thousands of years to be amplified and thousands of years to be amplified and studiedstudied

How does PCR work?How does PCR work?Need: Need: • The DNA molecule you The DNA molecule you

want to copy ( the want to copy ( the template)template)

• 2 DNA primer 2 DNA primer molecules – short, molecules – short, single stranded chains single stranded chains of nucleotides that of nucleotides that attach to the “unzipped attach to the “unzipped DNA” DNA”

• DNA polymeraseDNA polymerase• A system for heating A system for heating

and cooling the DNA and cooling the DNA strandsstrands

The Steps…..The Steps….. 1. 1. DNA molecule separated by DNA molecule separated by heatingheating the sample the sample

to 90-96 °Cto 90-96 °C

2. Strands are cooled so that 2. Strands are cooled so that DNA primersDNA primers can be can be added. Primers bind to complementary bases on added. Primers bind to complementary bases on the two separated strands of DNA.the two separated strands of DNA.

3. A special DNA polymerase (3. A special DNA polymerase (Taq polymeraseTaq polymerase) ) reads the template strand and matches it with reads the template strand and matches it with complementary nucleotides found within the DNA complementary nucleotides found within the DNA suspension (test tube) suspension (test tube)

Note: Taq polymerase comes from Note: Taq polymerase comes from Thermus aquaticusThermus aquaticus – a type of bacteria that lives in hot springs– a type of bacteria that lives in hot springs

END RESULT – 2 new helixes each made up of one of END RESULT – 2 new helixes each made up of one of the original strands plus a newly assembled the original strands plus a newly assembled complementary strandcomplementary strand

Each subsequent cycle doubles the # DNA molecules Each subsequent cycle doubles the # DNA molecules copied – exponential increasecopied – exponential increaseEx: 30 cycles – 2Ex: 30 cycles – 23030 DNA copied DNA copied

Each cycle takes 1-3 minutes. Therefore, in 1 hour you Each cycle takes 1-3 minutes. Therefore, in 1 hour you can have over a billion strands of a specific DNA can have over a billion strands of a specific DNA moleculemolecule