From Haystacks to Needles AP Biology Fall 2010
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
Gene Library
Isolating Genes A particular gene of interest can be isolated from other genes by use of probes
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
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
Big Time Amplification – PCR The polymerase chain reaction (PCR) can be used to make millions of copies of cDNA
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
Big Time Amplification – PCR What are the reaction steps? 1. Researchers mix primers, DNA polymerase, cellular DNA from an organism, and free nucleotides
Big Time Amplification – PCR 2. Precise temperature cycles cause the DNA strands to separate, exposing the bases
Big Time Amplification – PCR 3. Primers become positioned on the exposed nucleotides to form new copies of the original DNA
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