BIO 101 1st EditionLecture 20Outline of Last LectureI. Chromosomal basis of sexII. Genetics Problemsa. X-linked diseasesb. Chromosomal alterationsi. Change in Chromosome numberii. Changes in the structure of the chromosomeOutline of Current LectureI. DNA as the genetic materiala. Transformation Experimentsb. Bacteriophage experimentsII. DNA StructureIII. DNA ReplicationChapter 16- Search for the Genetic Material1940- known that chromosomes carry the genetic information and that chromosomes are made of DNA and protein. Most scientists thought that PROTEIN was the genetic material.Two Lines of Evidence that DNA is actually the genetic material, NOT PROTEIN:1. Transformation Experiments- means take up of external genetic information by the cell1928- Frederick Griffith did transformation experiments with bacteria2 strains of bacteria were used: S Strain (smooth), R strain (rough)- Grew S strain bacteria and then killed them and broke open the cells“S Soup” which contained everything in the cell, including the genetic material- Put R strain bacteria into “S Soup”- Some R bacteria took up genetic material and acquired the S phenotype= transformation1940- Avery and colleagues spent 10 years to identify the “transforming chemical in “S Soup””- it was DNA (still not accepted by most people at that time)2. Bacteriophage experiments- virus that infects bacteria- Virus is made of nucleic acid (DNA or RNA) and protein- it gets into a cell and makes more virus (i.e. it carries genetic information)- Is the genetic info in viruses nucleic acid or protein?- 1952- Alfred Hershey and Margaret Chase experiment- used a bacteriophage called T2 (made of DNA and protein)- They made 2 kinds of T2- 1) T2 with radioactive protein, 2) T2 with radioactive DNA- Found only DNA gets into the bacteria- Therefore, DNA not PROTEIN must be genetic materialBIO 101 1st EditionDNA Structure Review- DNA has 2 sugar-phosphate chains- Each sugar has one of 4 nitrogenous bases covalently attached to it (A, T, C, and G)- The two sugar-phosphate chains are held together by hydrogen bonds between the nitrogenous bases on one strand and the nitrogenous bases on the other strand- The hydrogen bonds are always A:::T (2 hydrogen bonds) or C:::G (3 hydrogen bonds)- Each sugar-phosphate chain has one 5’ end and one 3’ endo The 5’ end has a free phosphate on carbon #5 of the sugaro The 3’ end has a free –OH on carbon #3 of the sugar- The overall structure of the molecule is like a ladder- The 2 sugar-phosphates run in opposite directions called ANTI-PARALLEL- The hydrogen bonded nitrogenous bases are like the rungs of the ladder- The whole molecules is then twisted into a helix (spiral)o Sugar phosphate chains are on the outside of the moleculeo The hydrogen bonded bases are on the inside of the molecule- The 2 strands are said to be COMPLEMENTARY to each other (paired up by base pairing rules (C:::G and A:::T))Semi-Conservative DNA Replication Overview1. Starts at special places on the DNA called “origins of replication”, bacteria have 1 origin, eukaryotes have 100’s to 1000’s of origins2. The 2 strands of DNA come apart like a zipper at the H-bonded nitrogenous bases. This requires 3 proteins- Helicase- unwind the spiral DNA- Single-stranded binding protein- keeps 2 strands apart- Topoisomerases- untangle kinks3. Each strand acts as a template to make a new complementary strand (by the base pairing rules)4. Nucleotides (building blocks for DNA) for the new strand line up along the template strand (bases pairing rules C:::G and A:::T)BIO 101 1st Edition5. An enzyme called DNA Polymerase III hooks the lined up nucleotides together- Process requires a small RNA primer and a protein called primase- New nucleotides are added to this RNA primer only in 5’ 3’ direction(i.e. can ONLY add a nucleotide to an existing 3’-OH)- The Leading Strand is made as a single polymer moving toward the replication fork- The Lagging Strand- made by more complicated process- Has to be made in short pieces called Okazaki Fragments- because it goes in opposite direction from replication fork - Each Okazaki fragment starts with an RNA primers- RNA primers are removed and replaced with DNA by DNA POLYMERASE I- The DNA pieces are hooked together by DNA LigaseSemi-Conservative Replication= each new molecules has one strand from parent molecule and one newly made strandSemi-Conservative DNA Replication- Complex process- highly regulated- Is very rapid (~500 nucleotides added per second)- Is accurateo Initially= 1 base pair mistake per 10,000o Proof reading= 1 base pair mistake per 1
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