1. From Haystacks to Needles AP Biology Fall 2010

2. Isolating Genes  Gene library: a collection of bacteria that house different cloned DNA fragments, one of which is of interest May be the entire genome or of cDNA, which is free of introns

3. Gene Library

4. Isolating Genes  A particular gene of interest can be isolated from other genes by use of probes

5. Isolating Genes  DNA probes: short DNA sequences assembled from radioactive nucleotides Can pair with parts of the gene to be studied  This nucleic acid hybridization technique can be used with other procedures to select cells and their DNA May be of interest to a researcher

6. Isolating Genes  If researchers know the desired gene sequence, they use it to construct a primer by synthesizing the single stranded DNA  If the sequence is unknown, they use DNA from the same gene of a closley related species that has already been isolated

7. Big Time Amplification – PCR  The polymerase chain reaction (PCR) can be used to make millions of copies of cDNA

8. Big Time Amplification – PCR  What is needed for the reaction? Primers are short nucleotide sequences that are made in the laboratory They are recognized by DNA polymerases as the START tags for building complementary sequences of DNA dictated by computer programs stored in the machines

9. Big Time Amplification – PCR  What are the reaction steps? 1. Researchers mix primers, DNA polymerase, cellular DNA from an organism, and free nucleotides

10. Big Time Amplification – PCR 2. Precise temperature cycles cause the DNA strands to separate, exposing the bases

11. Big Time Amplification – PCR 3. Primers become positioned on the exposed nucleotides to form new copies of the original DNA

12. Big Time Amplification – PCR 4. Each round of reactions doubles the number of DNA molecules to eventually produce billions of molecules from very tiny amounts of original DNA http://www.youtube.com/watch?v=_YgXcJ4n-kQ