PCR-MichiganStateUniversity.pptVIP

Polymerase Chain Reaction(PCR) PCR is a means to amplify a particular piece of DNA Amplify= making numerous copies of a segment of DNA PCR can make billions of copies of a target sequence of DNA in a few hours PCR was invented in the 1984 as a way to make numerous copies of DNA fragments in the laboratory Its applications are vast and PCR is now an integral part of Molecular Biology DNA Replication vs. PCR PCR is a laboratory version of DNA Replication in cells The laboratory version is commonly called “in vitro” since it occurs in a test tube while “in vivo” signifies occurring in a living cell. DNA Replication in Cells (in vivo) DNA replication is the copying of DNA It typically takes a cell just a few hours to copy all of its DNA DNA replication is semi-conservative (i.e. one strand of the DNA is used as the template for the growth of a new DNA strand) This process occurs with very few errors (on average there is one error per 1 billion nucleotides copied) More than a dozen enzymes and proteins participate in DNA replication Key enzymes involved in DNA Replication DNA Polymerase DNA Ligase Primase Helicase Topoisomerase Single strand binding protein DNA Replication enzymes: DNA Polyerase catalyzes the elongation of DNA by adding nucleoside triphosphates to the 3’ end of the growing strand A nucleotide triphosphate is a 1 sugar + 1 base + 3 phosphates When a nucleoside triphosphate joins the DNA strand, two phosphates are removed. DNA polymerase can only add nucleotides to 3’ end of growing strand Complementary Base-Pairing in DNA DNA is a double helix, made up of nucleotides, with a sugar-phosphate backbone on the outside of the helix. Note: a nucleotide is a sugar + phosphate + nitrogenous base The two strands of DNA are held together by pairs of nitrogenous bases that are attached to each other via hydrogen bonds. The nitrogenous base adenine will only pair with thymine The nitrogenous base guanine will only pair with cytosine During replication, once t