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Molecular Genetics “Chapter 20” By the 1950s scientists had determined that chromosomes contained DNA and that DNA was the genetic material. The goal of molecular genetics is to find out how DNA determines phenotype of an organism. What happens to DNA during duplication of chromosomes in mitosis? Questions Questions 1-4 page 660 Exploration page 661 The Importance of DNA Deoxyribonuceic Acid (DNA) – is the carrier of genetic information in cells the nucleus of every cell in you body contain DNA. This molecule is found in the cells of all organisms, mushrooms to mammals. Only molecule known to replicate itself. (permits cell division) Guides the repair of worn cells parts and construction of new ones Provides for continuity of life Accounts for diversity of life forms Regulate the production of cell protein Genes – are the units of heredity in the chromosome, hair colour, skin colour, nose length. Genes are packed within DNA to make you unique. Unless you are an identical twin you DNA code is one of a kind Continuity of Life – is a succession of offspring that share structural similarities with those of their parents. Mutations – arise when the DNA within a chromosome is altered. Most mutations change the appearance of the organism Why do we study DNA? We study DNA for many reasons, e.g., its central importance to all life on Earth, medical benefits such as cures for diseases, better food crops. DNA DNA is often called the blueprint of life. In simple terms, DNA contains the instructions for making proteins within the cell. Chromosomes and DNA Our genes are on our chromosomes. Chromosomes are made up of a chemical called DNA. Historical Profile: James Watson and Francis Crick Used X –ray diffraction to help determine the structure of DNA molecule. Compared DNA structures in different organisms DNA consists of two strands of nucleotides. Each nucleotide contains a deoxyribose sugar, a phosphate group and a nitrogenous base. Knew that DNA was made up of sugars, phosphates and four different nitrogen bases: adenine, guanine, cytosine and thymine. Did not know how these bases were arranged. Did show that the proportion of these nitrogen bases varied in species, but the proportion stays the same in the DNA of all species. Ex: number of adenine is the same as thymine, guanine is the same as cytosine. This suggested that the nitrogen bases were arranged in pairs. If DNA in your body were stretched out it would reach to the sun and back appr. 3000 times. DNA Structure Watson and Crick’s Model also indicated that the two strands of DNA form a structure that resembles a twisted ladder. The base pairs are the rungs of the ladder and the sugar-phosphates backbones are the struts. This structure is called a double helix Each DNA strand in the double helix twists in a clockwise direction. DNA Strand In the DNA molecule, the bases from one strand are paired with bases in the other strand. Purine is always paired with pyrimidine, Ex: Adenine (purine) + thymine (pyrimidine) guanine (purine) + cytosine (pyrimidine) This type of pairing is termed complemetary base pairing. Hydrogen bonds between the complementary bases on opposite strands hold the double helix together. Although hydrogen bonds are weak, large numbers of hydrogen bonds are collectively strong, so the DNA molecule is very stable. Diagram Adenine forms two hydrogen bonds with thymine, while guanine forms three hydrogen bonds with cytosine. 5’ – ATGCCGTTA – 3’ 3’ – TACGGCAAT – 5’ The two strands of nucleotides are antiparallel (parallel but in opposite directions). One strand will have a 5’ carbon phosphate group at one end and a 3’ carbon and hydroxyl group of a deoxyribose sugar at its other end. Questions Questions 1-3 page 663 Activity Structure of DNA Review Described as a double helix. Resembles a twisted ladder. Sugar and phosphate molecules form the back bone of the ladder. Nitrogen bases form the rungs. Nitrogen bases from one spine of the ladder are connected with nitrogen bases from the other spine By means of hydrogen bonds a hydrogen bond is a weak bond that forms between that positive charge in the end of one molecule and a negative charge on the end of another molecule The backbone of DNA becomes twisted (spiral staircase) The DNA molecule is made up of individual units composed of deoxyribose sugars, phosphates, and nitrogen bases. Each unit is referred to as a nucleotide. NOTE: adenine pairs with thymine and Guanine pairs with cytosine The Shape of the Molecule DNA is a very long polymer. The basic shape is like a twisted ladder or zipper. This is called a double helix. The Double Helix Molecule The DNA double helix has two strands twisted together. One Strand of DNA The backbone of the molecule is alternating phosphates and deoxyribose sugar The teeth are nitrogenous bases. phosphate deoxyribose bases O O -P O Nucleotides O O O -P O O One deoxyribose together with its phosphate and base make a nucleotide . O O -P O O Phosphate Nitrogenous base C O C C C C O Deoxyribose One Strand of DNA One strand of DNA is a polymer of nucleotides. One strand of DNA has many millions of nucleotides. nucleotide Searching for the Chemical of Heredity Chromosomes – are the long threads of genetic material found in the nucleus of cells. Chromosomes are composed of many nucleic acids and proteins. Proteins are composed of 20 different amino acids, which can be organized in an infinite variety. Nucleotides – are the building blocks of nucleic acids. Nucleotides are composed of a ribose sugar, phosphate, and a nitrogen base. Four nitrogenous bases DNA has four different bases: Cytosine C Thymine T Adenine A Guanine G Two Kinds of Bases in DNA Pyrimidines are single ring bases. N N C O C C N C Purines are double ring bases. N N C C C N N C N C Thymine and Cytosine are pyrimidines Thymine and cytosine each have one ring of carbon and nitrogen atoms. N O C C O C C N C thymine N O C C N C N C cytosine Adenine and Guanine are purines Adenine and guanine each have two rings of carbon and nitrogen atoms. N C Adenine N C C N O N C N N C N C C C N Guanine C N N C Two Stranded DNA Remember, DNA has two strands that fit together something like a zipper. The teeth are the nitrogenous bases but why do they stick together? C N N C N C C C C N N N C C C O The bases attract each other because of hydrogen bonds. Hydrogen bonds are weak but there are millions and millions of them in a single molecule of DNA. The bonds between cytosine and guanine are shown here with dotted lines N Hydrogen Bonds N O Hydrogen Bonds, cont. When making hydrogen bonds, cytosine always pairs up with guanine N Adenine always pairs up with C thymine Adenine is bonded to thymine here N N O C N C C C N C C N N C C O C Chargraff’s Rule: Adenine and Thymine always join together A T Cytosine and Guanine always join together C G DNA by the Numbers Each cell has about 2 m of DNA. The average human has 75 trillion cells. The average human has enough DNA to go from the earth to the sun more than 400 times. The earth is 150 billion m or 93 million miles from DNA has a diameter of the sun. only 0.000000002 m. DNA Replication For mitosis to occur, DNA must copy itself and be equally divided between daughter cells. DNA replication – is the process by which a cell makes an exact copy of its DNA. DNA replication is semiconservative. Semiconservative replication – involves separating the two parent strands and using them to synthesis two new strands. The hydrogen bonds between complementary bases break, allowing the DNA helix to unzip. Each single DNA strand acts as a template to build the complementary strand. DNA replicating Semiconservatively Each daughter molecule receives one strand from the parent molecule plus one newly synthesized strand. Separating the DNA Strands The enzyme DNA helicase unwinds the helix by breaking the hydrogen bonds between the complementary bases. Proteins bind to the separated DNA strands to prevent them from reforming. The point at which the two strands are separating is called the replication fork. Separating DNA Strands Building the Complementary Strands Need to synthesis two new DNA strands on the template strands through complementary base pairing. New strands are sythesized by an enzyme called DNA polymerase III . This DNA polymerase builds a new strand by linking together free nucleotides that have bases complementary to bases in the template A short piece of single stranded ribonucleic acid, called a primer, is attached to the template strand. This gives DNA polymerase II a starting point to begin synthesizing a new DNA Strand. DNA polymerase III adds nucleotides to a growing strand in only one direction the 5’ – 3’ direction. This strand is called the leading strand. (synthesized towards the replication fork) The other new strand is synthesized in short fragments away from the replication fork. This is called the lagging strand. Another enzyme DNA ligase links the sugarphosphate backbone of the DNA fragments together. Building the Lagging Strand DNA Repair Complementary strands of DNA are synthesized, both DNA polymerase I and III act as quality control checkers by proofreading the newly synthesized strands. When a mistake occurs, they backtrack to the incorrect nucleotide, cut it out and continue adding. The repair must be made immediately to avoid the mistake from being copied in later replications. Questions Questions 1- 7 page 666. Gene Expression Specific segments on DNA are called genes Genes determine inherited traits in an organism The way the information in a gene is converted into a specific characteristic or trait through the production of a polypeptide is called geneexpression. Remember that proteins form many structures in an organism A second type of nucleic acid is involved in converting the instructions in a gene into a polypeptide chain. Ribonucleic acid (RNA) is a polymer of nucleotides similar tmin eo DNA. There are 3 main structural difference between RNA and DNA. First the sugar in RNA has an extra hydroxyl group and is called ribose. Second instead of the base thymine it has uracil. Third, RNA is a single stranded Central Dogma Two stages in gene expression: 1) transcription 2) translation Transcription the genetic information is converted from a DNA sequence into messenger mRNA. In all cells, the mRNA carries genetic information from the chromosomes to the site of protein synthesis. Translation the genetic information carried by mRNA is used to synthesize a polypeptide chain. This two step process of transferring information from DNA to RNA and then from RNA to protein is known as the central dogma of molecular genetics EX: DNA RNS ATGCAA UACGUU The Central Dogma of Molecular Genetics Transcription The DNA sequence of a gene is copied (transcribed) into the sequence of a single stranded mRNA molecule. Transcription is divided into 3 processes: 1) initiation 2) elongation 3) termination Initiation – an enzyme called RNA polymerase binds to the DNA at a specific site near the beginning of the gene. Elongation – RNA polymerase uses the DNA as a template to build the mRNA molecule. Termination – the RNA polymerase passes the end of the gene and comes to a stop. Summary of the Process of Transcription Question Questions 1-2 page 669 Translation Involves protein synthesis Only 20 amino acids are found in proteins Codon sequence of three bases in DNA Start codon (AUG) that signals the start of translation Stop codons (UAA, UAG, UGA) that signal the end of translation. Translation can be divided into the same three stages: initiation, elongation, termination. Translation Initiation – occurs when a ribosome recognizes a specific sequence on the mRNA and binds to that sight. The ribosome then moves along the mRNA in the 5’ – 3’ direction adding a new amino acid to the growing polypeptide chain each time it reads a codon. Ribosomes synthesize different proteins by associating with different mRNAs and reading their sequences. A ribosome must begin reading in the correct place. (Start codon) Once the ribosome has bound the mRNA, how does it get the amino acids that correspond to the codon? Second type of RNA molecule known as transfer RNA (tRNA). tRNA has a sequence of three bases, the anticodon that is complementary to the codon of the mRNA. Questions Questions 3-5 page 671 Elongation – The process is similar to a ticker tape running through a ticker tape machine. Start codon is AUG. The AUG codon also codes for methionine, so every protein initially starts with the amino acid methionine. The ribosome has two sites for tRNA to attach: A site and the P site. The tRNA with the anticodon complementary to the start codon enters the P sight. The next tRNA carrying the required amino acids enters the A site A peptide bond has formed between the methionine ans the second amino acid alanine. The ribosome has shifted over one codon so that the second tRNA is now in the P site. Refer to the diagram on the next page. Protein Synthesis Termination – eventually the ribosome reaches one of the 3 stop codons: UGA UAG, UAA. Since they do not code for an amino acid, there are no corresponding tRNAs Questions Questions 1-11 page 676 DNA and Biotechnology Recombinant DNA – fragment of DNA composed of sequences originating from at least two different sources. Ex: Gene that encodes insulin has been introduced into bacterial cells so that they become living factories producing this vital hormone. Genetic transformation- the introduction and expression of foreign DNA in an organism. DNA Sequencing DNA can be sequenced in a test tube using isolated segments of DNA. This technology requires a primer, DNA polymerase and 4 DNA nucleotides. The Human Genome Project Plans to produce maps of the entire genetic makeup of a human being. Human genome consists of approx. 30 000 genes Goal to help combat genetic disorders. The project also opened a box of ethical questions, legal dilemmas and societal problems. Enzymes and Recombinant DNA Anther way to identify specific segments of DNA is by creating genetic linkage maps. Once a segment of DNA has been identified, molecular biologists may use enzymes to isolate that segment or modify it. The DNA fragment may then be used to create recombinant DNA or be transferred to another organism. Restriction Endonucleases Known as restriction enzymes, are like molecule scissors that can cut double stranded DNA at specific base –pair sequences. Each type recognizes a particular sequence of nucleotides that is known as its recognition site. Molecular biologists use these enzymes to cut DNA in a predictable and precise way. Methylases Are enzymes that modify a restriction enzyme recognition site by adding a methyl group to one of the bases in the site. Important tools in recombinant DNA. They protect the gene fragment from being cut in the wrong spot. DNA Ligase To create recombinant DNA, pieces of DNA from two sources must be joined. Using restrictive enzymes and methylases, molecular biologists can engineer fragments of DNA that they want. These segments of DNA are joined together by DNA ligase. Transformation Is any process by which foreign DNA is incorporated into the genome of the cell. Vector – is the delivery system used to move the foreign DNA into a cell. Transgenic – a cell or an organism that is transformed by DNA from another species. Transformation of Bacteria Most common to be transformed. Used to study gene transformations, and gene functions First step is to isolate the DNA fragment to be tranferred. DNA is then introduced to a vector. Plamids are small circular double stranded DNA molecules that occur naturally in the cytoplasm of bacteria. Can be used as a vector. Refer to page 684 Figure 9 Questions Questions 1-9 page 686 Mutations and Genetic Variation Mutations are changes in the sequence of DNA. Are sources of new genetic variation Point mutations – are changes in a single base pair of a DNA sequence. They may or may not change the sequence of amino acids. Gene Mutations – change the amino acids specified by the DNA sequence, and they often involve more than a single base pair. Different Types of Mutations One type of point mutation, called a silent mutation, has no effect on the operation of the cell. ACA changed to ACU both code for threonine. Missense mutation – arises when a change in the base sequence if DNA alters a codon leadign to a different amino acid Ex: sickle cell anemia Nonsense mutation – occurs when a change in the DNA causes a stop codon. Deletion – is a mutation which occurs when nucleotides are removed from the DNA sequence. Insertion – is the placement if an extra nucleotide in a DNA sequence. Frameshift mutation – a mutation that causes the reading frame of codons to change. Translocation – the transfer of a fragment of DNA from one site in the genome to another location. Inversion – the reversal of a segment of DNA within a chromosome. Causes of Genetic Mutations Some are simply caused by error of genetic machinery and are known as spontaneous mutations. Ex: DNA polymerase misses a base or two causing point mutations. Mutations might arise from mutagenic agents. These are induced mutations Ex: UV radiation, cosmic rays, X-rays, chemicals. Case Study Case study page 689-690 Questions Questions 1-10 page 694 Review questions page 702-709 Homework Case Study – page 518-519 (old text) Homework Review Questions Page 522 Questions 1-8