Reading Guide Chapter 8: Bacterial Genetics This chapter is a continuation of the genetics theme with a focus on the different kinds of mutations that can occur, what can promote mutations, how to repair/fix mutations, and how to select for mutants or use bacteria to study these mutations. We will focus on sections 8.1-8.5 for now. We will finish the last sections, 8.6-8.9, after talking about viruses. So let’s begin with a look at some key terms and the different types of mutations that can occur in bacterial cells. Bacterial cells are good models to use for genetic research since they are haploid, or have one copy of every gene. One copy of every gene makes it easy to study the phenotype, or observable characteristics. Genetic changes can occur as a result of two main things, mutations and gene transfer. When a change in the genotype occurs and results in a mutation, this change is transferred to other cells in vertical gene transfer. If the change occurs as a result of gene transfer, this is called horizontal gene transfer. A few other key terms to review are wild-type and auxotroph. A wild-type cell refers to an organism that was isolated in nature, where an auxotroph refers to a cell that is a mutant. Often this mutant term is also connected to the cell lacking the ability to grow without a particular nutrient available. For example, E. coli can normally grow fine on a GSA plate generating all of the necessary growth factors from glucose. If this organism (the wild-type) is mutated and the results are an organism that lacks the ability to produce the amino acid histidine, then this is now considered to be an auxotroph lacking the ability to produce histidine. Mutations in bacterial cells are often the result of spontaneous events and it is through these mutations that organisms can respond to changes in their environment and natural selection occurs. Spontaneous mutations can occur as a result of errors with DNA polymerase, for example if the enzyme does not properly proofread a miss-paired base. They can also occur if cells are exposed to UV light which promotes the formation of thymine dimers in the DNA. If a mutation occurs which changes the sequence of DNA by just one base, this is called a base substitution or point mutation. The results of this mutation can be a silent mutation (no change in the amino acid code), missense mutation (a change in the code which results in a new amino acid) and a nonsense mutation (a change in the code which results in a stop codon). Some mutations are the result of the addition or deletion of one or more bases, this shifts the reading frame (the order in which you read the codons). This is known as a frameshift mutation. There are many different things that can promote mutation in cells. In my lecture I focus on three different types of mutagens or things that can cause mutations. They are chemicals such as nitrous acid, uv light, and transposons also known as jumping genes. Transposons were first discovered by Barbara McClintock as she studied the variegated patterns in corn. She proposed that the changes in the corn were due to the inactivation of genes for color of the actual kernels of corn. She was right! These changes in color are due to mobile pieces of DNA that randomly insert into existing DNA. If they happen to insert in to a coded region for a protein, the protein will be inactivated or no longer be made. Today we recognize that there are transposons found to infect all types of cells from plants to animals. Chemicals can cause mutations and one that is well studied is nitrous acid. This chemical alters the base adenine causing it to be mispaired with the base cytosine instead of pairing with the base thymine. Some chemicals are considered to be base analogs since the chemical structure of the compound is similar to the actual base. When the chemical is inserted into the DNA strand instead of the regular base, they will promote the wrong base pairing to occur. Intercalating agents are chemicals which insert into the DNA structure and physically cause an addition of a base when the DNA is replicated. Radiation can also cause mutations in cells. We have seen the effects of UV light on bacterial cells in lab. The UV light promotes the formation of thymine dimers which changes the shape of the structure of the DNA. This change in shape prevents DNA replication as well as protein synthesis unless repaired. X rays cause serious damage to DNA in the form of double strand breaks. These changes to the DNA strand cannot be repaired. Which brings me to the last topic for this reading guide…the topic of repair of damaged DNA. Bacterial cells can repair wrong base pairing by either the proof reading feature of DNA polymerase or by a process called mismatch repair. Mismatch repair requires an endonuclease to cut the DNA strand around the area where the wrong base is inserted. This leaves a gap in the strand of DNA which can be filled in by DNA polymerase and finally the fragments can be bonded to each other with the help of the enzyme DNA ligase. Note that these are the same key enzymes that were needed to synthesize the lagging strand of DNA in DNA replication. Repair of thymine dimers caused by uv damage can occur either in the presence of light or in the dark. The light repair mechanism is much simpler as it requires one enzyme (photolyase enzyme) that works in the light to break the thymine dimer bond. Dark repair requires an enzyme that recognizes the thymine dimer structure and cut the DNA strand. The resulting gap in the DNA is filled in by DNA polymerase, the pieces joined by DNA ligase, and you are done! SOS repair is the last mechanism to mention. In this type of repair the cell will activate a DNA polymerase that is able to just “randomly insert” bases without reading a template. This mechanism works as a last ditch effort to save the cell (hence the name SOS) due to extensive uv damage when the other mechanisms mentioned above would not work. I’ll add more later for the last section in this chapter (8.5) on how to select for mutants.