Why are we using more dilute cultures for the killing curve while we are using the arg selection culture at full strength?
1. The reason that we are using a more dilute culture for the killing curve is because we want to know how UV light exposure at different time intervals effects that growth of yeast, however, the arg selection culture has to be at full strength because that is our control variable and we want an equal possible situation for yeast to be grow.
2.How would you find the arg+ mutants on the killing curve plates?
2. The way to find arg+ mutants on the killing curve plates is after each plate is exposed to their respective amount of time under the UV light we will count the number of mutations that grew on the plate and we will know which plate is viable for mutations to go from an arg- to arg+.
5.Why do we have to incubate the plates in the dark?
6. The reason that the plates have to be placed in a dark location after being exposed to the UV light is to prevent mutations happening to the colonies inside the plates. The formation of pyrimidine dimmers, the major type of damage caused by UV light, distorts the double helix and blocks transcription or replication past the damaged site. Direct reversal of DNA damage is one repair mechanism used to restore damaged DNA. This is the most energy efficient method.
6. How does our experimental design allow us to determine that the UV caused the reversion from arg- to arg+ and not simple spontaneous suppressor mutations?
7. The experimental design allows us to determine that the mutation arg- to arg+ is due to a revertant mutation. This is due because we are exposing the culture to UV light and this initiates the repair mechanisms to react and fix the DNA sequence to these colonies, we now if or if not this mechanism worked due to the number of colonies grown on the plate after a week
Exposure to UV light causes pyrimidine dimer formation. The pyrimidine dimer can be repaired by direct reversal and NER. It can also be bypassed by activation of the SOS response.
a. Which of these three pathways maintains the arginine auxotrophy? Why?
NER
b. Which of these three pathways changes the arginine auxotrophy to prototrophy? Why?
SOS
1. The reason we use more dilute cultures for the killing curve is to observe the effect of UV light exposure on the growth of yeast. By diluting the culture, we can better measure the impact of different time intervals of UV exposure. On the other hand, the arg selection culture needs to be at full strength to serve as the control variable. This ensures that we have a consistent and comparable baseline for yeast growth.
2. To find the arg+ mutants on the killing curve plates, we need to examine the number of mutations that grow on each plate after exposure to UV light. After each plate is exposed for its respective time interval, we count the colonies that have developed. By comparing the number of mutations on each plate, we can determine which plate is viable for mutations to occur from an arg- to an arg+ state.
3. The plates need to be incubated in the dark after exposure to UV light in order to prevent additional mutations from occurring in the colonies. UV light can cause pyrimidine dimers, a type of DNA damage that can distort the DNA structure and block transcription or replication. Incubating the plates in the dark minimizes the potential for additional DNA damage, allowing us to accurately assess the effects of UV light on the growth and mutation of the yeast colonies.
4. Our experimental design enables us to determine that the UV light caused the reversion from arg- to arg+. By exposing the cultures to UV light, we induce DNA damage in the form of pyrimidine dimers. This damage triggers the repair mechanisms of the yeast, and if successful, leads to the colonies reverting from arg- to arg+. We can assess the effectiveness of the repair mechanisms by counting the number of colonies that grow on the plates after a week. If there is a significant increase in the number of arg+ colonies compared to the control, it indicates that the UV light exposure caused the mutation reversion.
5. Among the three pathways mentioned - direct reversal, NER, and SOS response - the NER pathway is likely responsible for maintaining the arginine auxotrophy. NER (nucleotide excision repair) is a DNA repair mechanism that recognizes and removes pyrimidine dimers, which are caused by UV light exposure. Since the pyrimidine dimers are responsible for the arginine auxotrophy in this case, repair through NER would likely maintain the auxotrophy.
6. On the other hand, the SOS response pathway is responsible for changing the arginine auxotrophy to prototrophy. The SOS response is an emergency DNA repair pathway that is initiated when extensive DNA damage, such as pyrimidine dimers, is present. It promotes error-prone DNA synthesis to bypass the damaged sites, allowing for potentially more mutations. In the context of the arginine auxotrophy, the SOS response can lead to the reversion from arg- to arg+ by allowing for the occurrence of suppressor mutations that restore the ability to synthesize arginine.