Introduction Ch1 Exam 1 Study Guide A gene is a particular sequence of DNA that codes for a trait Genes are located on chromosome which are made up of DNA and proteins Alleles are different forms of the same gene i e brown eyes or green eyes Genotype is the actual DNA sequence in an organism Phenotype is the traits that they portray Structure of DNA Ch10 Three experiments led to the conclusion that DNA is the genetic material 1 Griffith s Assay to Transformation 2 Avery MacCleod and McCarty Griffith worked with Pheumococcus of two strains Smooth S and Rough R When the S strain was injected into mice its coat prevented the immune system from recognizing the virus and the mice died When the rough strain was injected into the mice it did not have this coat and therefore the immune system could recognize and destroy the strain allowing the mice to live Griffith then heat killed the S strain and injected it into the mice The mice lived However when Griffith mixed the heat killed S strain and the live R strain the mice died He did not prove that DNA held the genetic material but he did suggest that the R strain was being transformed to the S strain Worked with the same bacteria as Griffith They took the heat killed S strain and removed all lipids and proteins because they knew that genes consisted mostly of proteins and nucleic acids They separated the bacteria into three samples One sample was treated with protease to destroy the protein One sample was treated with ribonuclease to destroy RNA The third sample was treated with deoxyribonuclease to destroy DNA The treated samples were each added to the R strain Results showed that only the samples with DNA were transformed to the virulent S strain The sample with the destroyed DNA only contained the R strain They concluded that DNA is the transforming substance Other scientists however were not convinced Perhaps they did not kill the strains completely They thought that proteins held the genetic material because they have more variation with 20 amino acids whereas DNA only has 4 bases Used bacteriophages that inject their chromosomes into cells then form another generation of phages They grew two samples of phages one in radioactive sulfur 35S and the other in radioactive phosphorous 32P Sulfur is found only in protein and phosphorous is found only in DNA The treated samples were then allowed to infect E coli Then the protein coats of the ghost phages were separated from the cells It was found that radioactive sulfur was present in the protein coats of the cells but not in the next generation of phages The next generation of phages contained the radioactive phosphorous proving that DNA holds the genetic material 3 Hershey and Chase Purines have 2 rings adenine and guanine 1 2 Pyrimidines have 1 ring cytosine thymine and uracil Chargaff s Ratios the ratio of A T 1 and G C 1 but the ratio of A T G C varies among species Sugar phosphate backbone of DNA DNA strands are antiparallel 3 to 5 on one strand and 5 to 3 on the other DNA can have 1 of 3 different helical forms o A B right handed Put your right thumb up The direction that your fingers curl is the same direction that the double helix turns DNA is usually in the B form o Z left handed Pretty rare DNA vs RNA H bonded on the 2 carbon Thymine Double stranded usually OH bonded on the 2 carbon Uracil Single stranded usually DNA Replication and Recombination Ch12 Meselson Stahl Experiment proved that DNA replication is semi conservative Meselson and Stahl first grew E coli cells in a medium containing the isotope of nitrogen 15N so that all of the DNA contained 15N Then they transferred the bacteria to a medium containing lighter nitrogen 14N and allowed for DNA replication After one round of replication they centrifuged the DNA to measure by density The DNA formed one band in the tube which was inconsistent with the conservative model of replication which would have shown two bands one for the DNA with 15N and one for the DNA with 14N Meselson and Stahl allowed the E coli to replicate again in 14N and centrifuged again This time they found two bands of different densities which was consistent with the semi conservative model of replication Replication could not be dispersive because that would have produced only a single strand Requirements of DNA replication template of single stranded DNA deoxyribonucleoside triphosphates dNTPs base sugar phosphates DNA polymerase primers free 3 OH group to extend DNA is synthesized 5 3 Prokaryotes can replicate in one of two ways Theta model 3 Rolling circle model Replication fork DNA Polymerase I can synthesize DNA and remove RNA primers and replace them with DNA DNA Polymerase III synthesizes DNA and is much faster than DNA polymerase I 4 Transcription Prokaryotes Eukaryotes Promoter Sequence 35 consensus sequence TTGACA 10 TATA Box 35 consensus sequence 25 TATA Box The 100 base pairs upstream from the core promoter contain consensus sequences that are different for different genes that vary in both length and location The TATAA binding protein TBP and transcription factor IID TFIID are the first two things to bind in the core promoter Then additional proteins also bind in the core promoter and specific transcription factors bind to the regulatory promoter Enhancers can be 1000s of base pairs away upstream or downstream help initiate transcription Many proteins stay bound to recruit more RNA polymerases Not as tightly regulated downstream from the gene is a consensus sequence that cleaves the RNA from the DNA The polymerase continues to synthesize RNA past where we need it until the RAT1 protein comes in to knock it off Initiation The sigma factor a protein binds to the RNA polymerase forming the holoenzyme which can recognize and bind to the 35 consensus sequence and the 10 TATA box within promoter sequence The holoenzyme then begins to unwind the DNA strand Termination Rho independent mechanism relies on a particular sequence at the end of a gene that consists of a pair of inverted sequence of bases which forms a hairpin in the secondary structure of the mRNA which is like a speed bump After the hair pin a long sequence of A s Because A s form only 2 hydrogen bonds RNA polymerase is not tightly attached to the DNA and it will dissociate Rho dependent mechanism there is no long sequence of A s after the hairpin structure so the Rho protein acts as a bowling ball to knock the polymerase off the DNA strand Elongation RNA Polymerase starts adding