BCHM 3050 1st Edition Lecture 20 Outline of Last Lecture I. Steps in DNA Synthesisa. Initiationb. Elongation c. TerminationII. Structure of DNA Polymerase DimerIII. “Dimeric” Function of DNA PolymeraseIV. Overview of the E. coli “Replisome”V. Termination of DNA Replication in ProkaryotesVI. Ends of DNA are not completely replicatedVII. Telomere Structure and FunctionOutline of Current Lecture I. Telomerase FunctionII. Polymerase Chain Reaction (PCR)III. The “CR” in PCRCurrent LectureI. Telomerase Functiona. Telomeres recruit telomerase enzyme and telomerase brings in an RNA template in with it so that the DNA polymerase can fill in the rest of the nucleotidesb. Telomerase is a reverse transcriptase because it can synthesis its own RNAc. Telomerase provides the RNA primer and 3’ OH group for polymerase to fill in thegaps at the endd. Telomere repeats are only in the parent strande. Telomerase extends the 3’ end of the parents strand first, then uses the telomereas a template to fill in the top strandThese notes represent a detailed interpretation of the professor’s lecture. GradeBuddy is best used as a supplement to your own notes, not as a substitute.II. Polymerase Chain Reaction (PCR)a. 1980 – Dr. Kary Mullis decided to replicate DNA in test tube - PCRb. Make more than a billion in just 30 minutesc. Heat the DNA at 90-95 degrees C for 1-3 min to break the DNA strands apart à Denaturationd. This replaces the role of Helicase (without any origins, etc.)e. Primers bind to 3’ ends of separated strandsf. He added 2 primers, dNTPs (dATP/dGTP/dCTP/dTTP), buffer, DNA polymerase (Taq), template DNA in test tubeg. Had to decrease the temperature to allow the primers to join à Annealing (40-50 degrees C)h. Extension – 70 degrees C for about 5 mini. Denaturation mirrored Helicasej. Annealing mirrored RNA primase, ssBP, DNA polymerase IIIk. Extension mirrored DNA polymerase IIIIII. The “CR” in PCRa. Exponential amplification – 2nb. Primers always stick to 3’ end of parent strandc. Primers are usually 20-30 nucleotides longd. Possible exam questions: What would be the primers for this DNA sequence? (watch out for orientation of 5’ to
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