Evolution ExperimentsIntroduction | ||||||||
Changed: | ||||||||
< < | An evolution experiment consists of evolving one or more ancestor strains in laboratory culture and assessing the types and impacts of acquired mutations on the functions of that organism including its ability to reproduce, consume or produce compounds of interest, or resist particular stress conditions. The topics discussed on this page assume a general bacterial evolution experiment where a bacterial strain is serially passaged to fresh media over time, however there are other types of evolution experiments including evolving eukaryotes and viruses, passaging pathogens and symbionts between hosts, and evolving multiple strains in co-culture that require additional considerations. | |||||||
> > | An evolution experiment typically consists of evolving one or more ancestor strains in laboratory culture and assessing the types and impacts of acquired mutations on that organism's ability to reproduce, consume or produce compounds of interest, or resist particular stress conditions. The topics discussed on this page assume a general bacterial evolution experiment where a bacterial strain is serially passaged to fresh media over time, however there are other types of evolution experiments including evolving eukaryotes or viruses, passaging pathogens and symbionts between hosts, and evolving multiple strains in co-culture that require additional considerations. | |||||||
Changed: | ||||||||
< < | Broadly speaking there are two kinds of evolution experiments: adaptive evolution and mutation accumulation. In adaptive evolution experiments cells within a population compete with each other and generally trend towards increasing fitness. These experiments are useful for identifying mutations that improve a particular function, such as growth on a particular compound or resistance to a particular stress. Individuals with detrimental mutations are usually out-competed, thus the mutations evolved in these experiments are usually biased towards beneficial mutations. Mutation accumulation experiments, on the other hand, reduce or remove selection, allowing the persistence of all types of mutations and are thus useful for calculating overall mutation rates or examining the spectrum of possible mutations. In these experiments the effect of selection is usually limited by selecting colonies to streak from one plate to another at random. Since colonies are grown initially from individual cells this also represents the smallest possible bottleneck size, supporting genetic drift. The discussion below applies predominantly to adaptive evolution experiments, you can read more about mutation accumulation experiments here. | |||||||
> > | Broadly speaking there are two kinds of evolution experiments: adaptive evolution and mutation accumulation. In adaptive evolution experiments cells within a population compete with each other and generally trend towards increasing fitness. These experiments are useful for identifying mutations that improve a particular function, such as growth on a particular compound or resistance to a particular stress. Individuals with detrimental mutations are usually out-competed, thus the mutations evolved in these experiments are biased towards beneficial mutations. Mutation accumulation experiments, on the other hand, reduce or remove selection, allowing the persistence of all types of mutations and are thus useful for calculating overall mutation rates or examining the spectrum of possible mutations. In these experiments the effect of selection is usually limited by randomly selecting colonies to streak from one plate to another. Since colonies are grown initially from individual cells this also represents the smallest possible bottleneck size, supporting genetic drift. The discussion below applies predominantly to adaptive evolution experiments, you can read more about mutation accumulation experiments here. | |||||||
For an example protocol and overview of a specific adaptive evolution experiment see this paper describing the Long-term Evolution Experiment with E. coli.
Experimental designAn overall evolution experiment will likely consist of four phases: 1) inoculating populations, 2) routine transfers, 3) isolating clones and freezing stocks, and 4) analysis. 1) Inoculating populations Regardless of whether they are adaptive or mutation accumulation experiments, evolution experiments should be started by streaking the ancestral stock onto an agar plate and then initiating each population from a separate isolated colony. This reduces the impact of mutations that may already be present in the ancestral stock and ensures that the populations begin as single genotypes. It's a good idea to start more populations than you need in case some become contaminated later. Also set up a separate tube or flask with an uninoculated control to check for contamination. After the first day of growth populations should be frozen so they can be sequenced and referred to later, in particular when determining whether identical mutations shared between samples evolved during the experiment or were present in the initial culture. 2) Routine transfers Over the course of an evolution experiment the populations will be transferred to fresh media to continue growing after their initial media has been used up. Between transfers the pipette should be wiped with ethanol to prevent cross-contamination between samples. Designing an effective evolution experiment requires taking into account the impact of:
| ||||||||
Changed: | ||||||||
< < |
| |||||||
> > |
| |||||||
3) Isolating clones and freezing stocks At the end of the transfer period the evolved strains should be frozen for further analysis. This can be done as populations by freezing culture media from the last transfer and/or as isolates by streaking each population on a plate and inoculating a large, isolated colony into fresh media and then freezing it. Isolates can also taken from frozen populations after the experiment by thawing and streaking out the stock. | ||||||||
Changed: | ||||||||
< < | 4) Analysis The end result of an evolution experiment is likely isolated, evolved clones that have acquired additional mutations relative to their ancestor. The goals, then, are usually to identify these mutations in their genomes and determine what their functional consequences, if any, are.
| |||||||
> > | 4) Analysis The end result of an evolution experiment is likely isolated, evolved clones that have acquired additional mutations relative to their ancestor. The goals, then, are usually to identify these mutations in their genomes and determine what their functional consequences are, if any.
| |||||||
| ||||||||
Added: | ||||||||
> > |
ComplicationsSome potential difficulties that may arise during evolution experiments include:
|
Evolution ExperimentsIntroductionAn evolution experiment consists of evolving one or more ancestor strains in laboratory culture and assessing the types and impacts of acquired mutations on the functions of that organism including its ability to reproduce, consume or produce compounds of interest, or resist particular stress conditions. The topics discussed on this page assume a general bacterial evolution experiment where a bacterial strain is serially passaged to fresh media over time, however there are other types of evolution experiments including evolving eukaryotes and viruses, passaging pathogens and symbionts between hosts, and evolving multiple strains in co-culture that require additional considerations. | ||||||||
Changed: | ||||||||
< < | In general there are two kinds of evolution experiments: adaptive evolution and mutation accumulation. In adaptive evolution experiments cells within a population compete with each other and generally trend towards an increase in fitness. These experiments are useful for identifying mutations that improve a particular function, such as growth on a particular compound or resistance to a particular stress. Individuals with detrimental mutations are usually out-competed, thus the mutations evolved in these experiments are usually biased towards beneficial mutations. Mutation accumulation experiments, on the other hand, reduce or remove selection, allowing the persistence of all types of mutations and are thus useful for calculating overall mutation rates or examining the spectrum of possible mutations. In these experiments the effect of selection is usually limited by selecting colonies to streak from one plate to another at random. Since colonies are grown initially from individual cells this also represents the smallest possible bottleneck size, supporting genetic drift. The discussion below applies predominantly to adaptive evolution experiments, you can read more about mutation accumulation experiments here. | |||||||
> > | Broadly speaking there are two kinds of evolution experiments: adaptive evolution and mutation accumulation. In adaptive evolution experiments cells within a population compete with each other and generally trend towards increasing fitness. These experiments are useful for identifying mutations that improve a particular function, such as growth on a particular compound or resistance to a particular stress. Individuals with detrimental mutations are usually out-competed, thus the mutations evolved in these experiments are usually biased towards beneficial mutations. Mutation accumulation experiments, on the other hand, reduce or remove selection, allowing the persistence of all types of mutations and are thus useful for calculating overall mutation rates or examining the spectrum of possible mutations. In these experiments the effect of selection is usually limited by selecting colonies to streak from one plate to another at random. Since colonies are grown initially from individual cells this also represents the smallest possible bottleneck size, supporting genetic drift. The discussion below applies predominantly to adaptive evolution experiments, you can read more about mutation accumulation experiments here. | |||||||
For an example protocol and overview of a specific adaptive evolution experiment see this paper describing the Long-term Evolution Experiment with E. coli.
Experimental design | ||||||||
Changed: | ||||||||
< < | An overall evolution experiment will likely involve three phases: 1) inoculating populations, 2) routine transfers, and 3) isolating clones and freezing stocks. | |||||||
> > | An overall evolution experiment will likely consist of four phases: 1) inoculating populations, 2) routine transfers, 3) isolating clones and freezing stocks, and 4) analysis. | |||||||
1) Inoculating populations Regardless of whether they are adaptive or mutation accumulation experiments, evolution experiments should be started by streaking the ancestral stock onto an agar plate and then initiating each population from a separate isolated colony. This reduces the impact of mutations that may already be present in the ancestral stock and ensures that the populations begin as single genotypes. It's a good idea to start more populations than you need in case some become contaminated later. Also set up a separate tube or flask with an uninoculated control to check for contamination. After the first day of growth populations should be frozen so they can be sequenced and referred to later, in particular when determining whether identical mutations shared between samples evolved during the experiment or were present in the initial culture.
2) Routine transfers Over the course of an evolution experiment the populations will be transferred to fresh media to continue growing after their initial media has been used up. Between transfers the pipette should be wiped with ethanol to prevent cross-contamination between samples. Designing an effective evolution experiment requires taking into account the impact of:
| ||||||||
Changed: | ||||||||
< < |
| |||||||
> > |
| |||||||
| ||||||||
Changed: | ||||||||
< < | Analyzing evolution experimentsThe end result of an evolution experiment is likely isolated, evolved clones that have acquired additional mutations relative to their ancestor. The goals, then, are usually to identify these mutations in their genomes and determine what their functional consequences, if any, are. Mutations can be identified by sequencing the isolates' genomes and comparing them to the ancestor with breseq. Quantifying the impacts of the evolution is typically performed with fitness assays or other functional assays (i.e. growth curve, minimum inhibitory concentration, biochemical assays, as appropriate). Performing these assays on evolved isolates can confirm that the isolates have evolved the desired trait, however to tie the phenotype to a specific mutation the mutation should be reconstructed in the ancestor strain. These assays should be performed under the same conditions in which the populations were evolved (i.e. the same media) as the mutations may only be beneficial under those conditions. | |||||||
> > | 4) Analysis The end result of an evolution experiment is likely isolated, evolved clones that have acquired additional mutations relative to their ancestor. The goals, then, are usually to identify these mutations in their genomes and determine what their functional consequences, if any, are.
| |||||||
Added: | ||||||||
> > |
|
| ||||||||
Deleted: | ||||||||
< < | Page under construction (7/9/2024) | |||||||
Evolution ExperimentsIntroduction | ||||||||
Changed: | ||||||||
< < | An evolution experiment consists of evolving one or more ancestor strains in laboratory culture and assessing the types and impacts of acquired mutations on the functions of that organism including its ability to reproduce, consume or produce compounds of interest, or resist particular stress conditions. The topics discussed on this page assume a general bacterial evolution experiment where a bacterial strain is serially passaged to fresh media over time, however there are other types of evolution experiments including evolving phages and eukaryotes, passaging pathogens and symbionts between hosts, and evolving multiple strains in co-culture that require additional considerations. | |||||||
> > | An evolution experiment consists of evolving one or more ancestor strains in laboratory culture and assessing the types and impacts of acquired mutations on the functions of that organism including its ability to reproduce, consume or produce compounds of interest, or resist particular stress conditions. The topics discussed on this page assume a general bacterial evolution experiment where a bacterial strain is serially passaged to fresh media over time, however there are other types of evolution experiments including evolving eukaryotes and viruses, passaging pathogens and symbionts between hosts, and evolving multiple strains in co-culture that require additional considerations. | |||||||
In general there are two kinds of evolution experiments: adaptive evolution and mutation accumulation. In adaptive evolution experiments cells within a population compete with each other and generally trend towards an increase in fitness. These experiments are useful for identifying mutations that improve a particular function, such as growth on a particular compound or resistance to a particular stress. Individuals with detrimental mutations are usually out-competed, thus the mutations evolved in these experiments are usually biased towards beneficial mutations. Mutation accumulation experiments, on the other hand, reduce or remove selection, allowing the persistence of all types of mutations and are thus useful for calculating overall mutation rates or examining the spectrum of possible mutations. In these experiments the effect of selection is usually limited by selecting colonies to streak from one plate to another at random. Since colonies are grown initially from individual cells this also represents the smallest possible bottleneck size, supporting genetic drift. The discussion below applies predominantly to adaptive evolution experiments, you can read more about mutation accumulation experiments here.
For an example protocol and overview of a specific adaptive evolution experiment see this paper describing the Long-term Evolution Experiment with E. coli.
Experimental designAn overall evolution experiment will likely involve three phases: 1) inoculating populations, 2) routine transfers, and 3) isolating clones and freezing stocks. 1) Inoculating populations Regardless of whether they are adaptive or mutation accumulation experiments, evolution experiments should be started by streaking the ancestral stock onto an agar plate and then initiating each population from a separate isolated colony. This reduces the impact of mutations that may already be present in the ancestral stock and ensures that the populations begin as single genotypes. It's a good idea to start more populations than you need in case some become contaminated later. Also set up a separate tube or flask with an uninoculated control to check for contamination. After the first day of growth populations should be frozen so they can be sequenced and referred to later, in particular when determining whether identical mutations shared between samples evolved during the experiment or were present in the initial culture. 2) Routine transfers Over the course of an evolution experiment the populations will be transferred to fresh media to continue growing after their initial media has been used up. Between transfers the pipette should be wiped with ethanol to prevent cross-contamination between samples. Designing an effective evolution experiment requires taking into account the impact of:
| ||||||||
Changed: | ||||||||
< < |
| |||||||
> > |
| |||||||
Analyzing evolution experimentsThe end result of an evolution experiment is likely isolated, evolved clones that have acquired additional mutations relative to their ancestor. The goals, then, are usually to identify these mutations in their genomes and determine what their functional consequences, if any, are. Mutations can be identified by sequencing the isolates' genomes and comparing them to the ancestor with breseq. Quantifying the impacts of the evolution is typically performed with fitness assays or other functional assays (i.e. growth curve, minimum inhibitory concentration, biochemical assays, as appropriate). Performing these assays on evolved isolates can confirm that the isolates have evolved the desired trait, however to tie the phenotype to a specific mutation the mutation should be reconstructed in the ancestor strain. These assays should be performed under the same conditions in which the populations were evolved (i.e. the same media) as the mutations may only be beneficial under those conditions. |
Evolution ExperimentsIntroductionAn evolution experiment consists of evolving one or more ancestor strains in laboratory culture and assessing the types and impacts of acquired mutations on the functions of that organism including its ability to reproduce, consume or produce compounds of interest, or resist particular stress conditions. The topics discussed on this page assume a general bacterial evolution experiment where a bacterial strain is serially passaged to fresh media over time, however there are other types of evolution experiments including evolving phages and eukaryotes, passaging pathogens and symbionts between hosts, and evolving multiple strains in co-culture that require additional considerations. In general there are two kinds of evolution experiments: adaptive evolution and mutation accumulation. In adaptive evolution experiments cells within a population compete with each other and generally trend towards an increase in fitness. These experiments are useful for identifying mutations that improve a particular function, such as growth on a particular compound or resistance to a particular stress. Individuals with detrimental mutations are usually out-competed, thus the mutations evolved in these experiments are usually biased towards beneficial mutations. Mutation accumulation experiments, on the other hand, reduce or remove selection, allowing the persistence of all types of mutations and are thus useful for calculating overall mutation rates or examining the spectrum of possible mutations. In these experiments the effect of selection is usually limited by selecting colonies to streak from one plate to another at random. Since colonies are grown initially from individual cells this also represents the smallest possible bottleneck size, supporting genetic drift. The discussion below applies predominantly to adaptive evolution experiments, you can read more about mutation accumulation experiments here. | ||||||||
Added: | ||||||||
> > | For an example protocol and overview of a specific adaptive evolution experiment see this paper describing the Long-term Evolution Experiment with E. coli. | |||||||
Experimental designAn overall evolution experiment will likely involve three phases: 1) inoculating populations, 2) routine transfers, and 3) isolating clones and freezing stocks. 1) Inoculating populations Regardless of whether they are adaptive or mutation accumulation experiments, evolution experiments should be started by streaking the ancestral stock onto an agar plate and then initiating each population from a separate isolated colony. This reduces the impact of mutations that may already be present in the ancestral stock and ensures that the populations begin as single genotypes. It's a good idea to start more populations than you need in case some become contaminated later. Also set up a separate tube or flask with an uninoculated control to check for contamination. After the first day of growth populations should be frozen so they can be sequenced and referred to later, in particular when determining whether identical mutations shared between samples evolved during the experiment or were present in the initial culture. 2) Routine transfers Over the course of an evolution experiment the populations will be transferred to fresh media to continue growing after their initial media has been used up. Between transfers the pipette should be wiped with ethanol to prevent cross-contamination between samples. Designing an effective evolution experiment requires taking into account the impact of:
| ||||||||
Changed: | ||||||||
< < |
| |||||||
> > |
| |||||||
3) Isolating clones and freezing stocks At the end of the transfer period the evolved strains should be frozen for further analysis. This can be done as populations by freezing culture media from the last transfer and/or as isolates by streaking each population on a plate and inoculating a large, isolated colony into fresh media and then freezing it. Isolates can also taken from frozen populations after the experiment by thawing and streaking out the stock.
Analyzing evolution experiments | ||||||||
Changed: | ||||||||
< < | The end result of an evolution experiment is likely isolated, evolved clones that have acquired additional mutations relative to their ancestor. The goals, then, are usually to identify these mutations in their genome and determine what their functional consequences, if any, are. These can be accomplished by sequencing the isolates' genomes and comparing them to the ancestor with breseq and performing fitness assays or other functional assays (i.e. growth curve, minimum inhibitory concentration, biochemical assays, as appropriate), respectively. | |||||||
> > | The end result of an evolution experiment is likely isolated, evolved clones that have acquired additional mutations relative to their ancestor. The goals, then, are usually to identify these mutations in their genomes and determine what their functional consequences, if any, are. Mutations can be identified by sequencing the isolates' genomes and comparing them to the ancestor with breseq. Quantifying the impacts of the evolution is typically performed with fitness assays or other functional assays (i.e. growth curve, minimum inhibitory concentration, biochemical assays, as appropriate). Performing these assays on evolved isolates can confirm that the isolates have evolved the desired trait, however to tie the phenotype to a specific mutation the mutation should be reconstructed in the ancestor strain. These assays should be performed under the same conditions in which the populations were evolved (i.e. the same media) as the mutations may only be beneficial under those conditions. |
Evolution ExperimentsIntroduction | ||||||||
Changed: | ||||||||
< < | An evolution experiment consists of evolving one or more ancestor strains in laboratory culture and assessing the types and impacts of acquired mutations on the functions of that organism including its ability to reproduce, consume or produce compounds of interest, or resist particular stress conditions. The topics discussed on this page assume a general bacterial evolution experiment where a bacterial strain is serially passaged to fresh media over time. There are other types of evolution experiments including evolving phages, passaging pathogens and symbionts between hosts, and evolving multiple strains in co-culture that require additional considerations. | |||||||
> > | An evolution experiment consists of evolving one or more ancestor strains in laboratory culture and assessing the types and impacts of acquired mutations on the functions of that organism including its ability to reproduce, consume or produce compounds of interest, or resist particular stress conditions. The topics discussed on this page assume a general bacterial evolution experiment where a bacterial strain is serially passaged to fresh media over time, however there are other types of evolution experiments including evolving phages and eukaryotes, passaging pathogens and symbionts between hosts, and evolving multiple strains in co-culture that require additional considerations. | |||||||
Changed: | ||||||||
< < | In general there are two kinds of evolution experiments: adaptive evolution and mutation accumulation. In adaptive evolution experiments cells within a culture compete with each other and trend towards an increase in fitness. These experiments are useful for identifying mutations that improve a particular function, such as growth on a particular compound or resistance to a particular stress. Individuals with detrimental mutations are usually out-competed, thus the mutations evolved in these experiments is usually biased towards beneficial mutations. Mutation accumulation experiments, on the other hand, reduce or remove selection, allowing the persistence of all types of mutations and are thus useful for calculating overall mutation rates or examining the spectrum of possible mutations. The effect of selection is usually limited by selecting colonies to streak from one plate to another at randon. Since colonies are grown initially from individual cells this also represents the smallest possible bottleneck size. The discussion on this page applies predominantly to adaptive evolution experiments, you can read more about mutation accumulation experiments here. | |||||||
> > | In general there are two kinds of evolution experiments: adaptive evolution and mutation accumulation. In adaptive evolution experiments cells within a population compete with each other and generally trend towards an increase in fitness. These experiments are useful for identifying mutations that improve a particular function, such as growth on a particular compound or resistance to a particular stress. Individuals with detrimental mutations are usually out-competed, thus the mutations evolved in these experiments are usually biased towards beneficial mutations. Mutation accumulation experiments, on the other hand, reduce or remove selection, allowing the persistence of all types of mutations and are thus useful for calculating overall mutation rates or examining the spectrum of possible mutations. In these experiments the effect of selection is usually limited by selecting colonies to streak from one plate to another at random. Since colonies are grown initially from individual cells this also represents the smallest possible bottleneck size, supporting genetic drift. The discussion below applies predominantly to adaptive evolution experiments, you can read more about mutation accumulation experiments here. | |||||||
Experimental design | ||||||||
Changed: | ||||||||
< < | An overall evolution experiment will likely involve three phases: 1) inoculating populations, 2) routine transfers, and 3) isolating clones and freezing stocks. | |||||||
> > | An overall evolution experiment will likely involve three phases: 1) inoculating populations, 2) routine transfers, and 3) isolating clones and freezing stocks. | |||||||
Changed: | ||||||||
< < | 1) Inoculating populations Regardless of whether they are adaptive or mutation accumulation experiments, evolution experiments should be started by streaking the ancestral stock onto an agar plate and then initiating each population from a separate isolated colony. This reduces the impact of mutations that may already be present in the ancestral stock and ensures that the populations begin as single genotypes. Also set up a separate tube or flask with an uninoculated control to check for contamination. After the first day of growth populations should be frozen so they can be sequenced and referred to later, in particular when determining whether identical mutations shared between samples evolved during the experiment or were present in the initial culture. | |||||||
> > | 1) Inoculating populations Regardless of whether they are adaptive or mutation accumulation experiments, evolution experiments should be started by streaking the ancestral stock onto an agar plate and then initiating each population from a separate isolated colony. This reduces the impact of mutations that may already be present in the ancestral stock and ensures that the populations begin as single genotypes. It's a good idea to start more populations than you need in case some become contaminated later. Also set up a separate tube or flask with an uninoculated control to check for contamination. After the first day of growth populations should be frozen so they can be sequenced and referred to later, in particular when determining whether identical mutations shared between samples evolved during the experiment or were present in the initial culture. | |||||||
Changed: | ||||||||
< < | 2) Routine transfers Over the course of an evolution experiment the populations will be transferred to fresh media to continue growing after their initial media has been used up. Designing an effective evolution experiment requires taking into account the impact of:
| |||||||
> > | 2) Routine transfers Over the course of an evolution experiment the populations will be transferred to fresh media to continue growing after their initial media has been used up. Between transfers the pipette should be wiped with ethanol to prevent cross-contamination between samples. Designing an effective evolution experiment requires taking into account the impact of:
| |||||||
Changed: | ||||||||
< < | 3) Isolating clones and freezing stocks At the end of the transfer period the evolved strains should be frozen for further analysis. For an adaptive evolution experiment this can be done as populations by freezing culture media from the last transfer and/or as isolates by streaking each population on a plate and inoculating a large, isolated colony into fresh media and then freezing it. Isolates can also taken from frozen populations after the experiment by thawing and streaking out the stock. | |||||||
> > | 3) Isolating clones and freezing stocks At the end of the transfer period the evolved strains should be frozen for further analysis. This can be done as populations by freezing culture media from the last transfer and/or as isolates by streaking each population on a plate and inoculating a large, isolated colony into fresh media and then freezing it. Isolates can also taken from frozen populations after the experiment by thawing and streaking out the stock. | |||||||
Analyzing evolution experiments | ||||||||
Changed: | ||||||||
< < | The end result of an evolution experiment is likely isolated, evolved clones that have acquired additional mutations relative to their ancestor. The goals, then, are usually to identify these mutations in the genome and determine what their functional consequences, if any, are. These can be accomplished by sequencing the isolates' genomes and comparing them to the ancestor with breseq and performing fitness assays or other functional assays (i.e. growth curve, minimum inhibitory concentration, biochemical assays, as appropriate), respectively. | |||||||
> > | The end result of an evolution experiment is likely isolated, evolved clones that have acquired additional mutations relative to their ancestor. The goals, then, are usually to identify these mutations in their genome and determine what their functional consequences, if any, are. These can be accomplished by sequencing the isolates' genomes and comparing them to the ancestor with breseq and performing fitness assays or other functional assays (i.e. growth curve, minimum inhibitory concentration, biochemical assays, as appropriate), respectively. |
Evolution ExperimentsIntroductionAn evolution experiment consists of evolving one or more ancestor strains in laboratory culture and assessing the types and impacts of acquired mutations on the functions of that organism including its ability to reproduce, consume or produce compounds of interest, or resist particular stress conditions. The topics discussed on this page assume a general bacterial evolution experiment where a bacterial strain is serially passaged to fresh media over time. There are other types of evolution experiments including evolving phages, passaging pathogens and symbionts between hosts, and evolving multiple strains in co-culture that require additional considerations. | ||||||||
Changed: | ||||||||
< < | In general there are two kinds of evolution experiments: adaptive evolution and mutation accumulation. In adaptive evolution experiments cells within a culture compete with each other and trend towards an increase in fitness. These experiments are useful for identifying mutations that improve a particular function, such as growth on a particular compound or resistance to a particular stress. Individuals with detrimental mutations are usually out-competed, thus the mutations evolved in these experiments is usually biased towards beneficial mutations. Mutation accumulation experiments, on the other hand, reduce or remove selection, allowing the persistence of all types of mutations and are thus useful for calculating overall mutation rates or examining the spectrum of possible mutations. | |||||||
> > | In general there are two kinds of evolution experiments: adaptive evolution and mutation accumulation. In adaptive evolution experiments cells within a culture compete with each other and trend towards an increase in fitness. These experiments are useful for identifying mutations that improve a particular function, such as growth on a particular compound or resistance to a particular stress. Individuals with detrimental mutations are usually out-competed, thus the mutations evolved in these experiments is usually biased towards beneficial mutations. Mutation accumulation experiments, on the other hand, reduce or remove selection, allowing the persistence of all types of mutations and are thus useful for calculating overall mutation rates or examining the spectrum of possible mutations. The effect of selection is usually limited by selecting colonies to streak from one plate to another at randon. Since colonies are grown initially from individual cells this also represents the smallest possible bottleneck size. The discussion on this page applies predominantly to adaptive evolution experiments, you can read more about mutation accumulation experiments here. | |||||||
Experimental designAn overall evolution experiment will likely involve three phases: 1) inoculating populations, 2) routine transfers, and 3) isolating clones and freezing stocks. | ||||||||
Changed: | ||||||||
< < | 1) Inoculating populations Regardless of whether they are adaptive or mutation accumulation experiments, evolution experiments should be started by streaking the ancestral stock onto an agar plate and then initiating each population from a separate isolated colony. This reduces the impact of mutations that may already be present in the ancestral stock and ensures that the populations begin as single genotypes. After the first day of growth populations should be frozen so they can be sequenced and referred to later, in particular when determining whether identical mutations shared between samples evolved during the experiment or were present in the initial culture. | |||||||
> > | 1) Inoculating populations Regardless of whether they are adaptive or mutation accumulation experiments, evolution experiments should be started by streaking the ancestral stock onto an agar plate and then initiating each population from a separate isolated colony. This reduces the impact of mutations that may already be present in the ancestral stock and ensures that the populations begin as single genotypes. Also set up a separate tube or flask with an uninoculated control to check for contamination. After the first day of growth populations should be frozen so they can be sequenced and referred to later, in particular when determining whether identical mutations shared between samples evolved during the experiment or were present in the initial culture. | |||||||
2) Routine transfers Over the course of an evolution experiment the populations will be transferred to fresh media to continue growing after their initial media has been used up. Designing an effective evolution experiment requires taking into account the impact of: | ||||||||
Changed: | ||||||||
< < |
| |||||||
> > |
| |||||||
Added: | ||||||||
> > |
| |||||||
Changed: | ||||||||
< < | 3) Isolating clones and freezing stocks At the end of the transfer period the evolved strains should be frozen for further analysis. For an adaptive evolution experiment this can be done as populations by freezing a milliliter of culture from the last transfer and/or as isolates by streaking each population on a plate and inoculating a large, isolated colony into fresh media and then freezing it. Similarly, colonies from the last transfer of mutation accumulation experiments should be inoculated into cultures and frozen. | |||||||
> > | 3) Isolating clones and freezing stocks At the end of the transfer period the evolved strains should be frozen for further analysis. For an adaptive evolution experiment this can be done as populations by freezing culture media from the last transfer and/or as isolates by streaking each population on a plate and inoculating a large, isolated colony into fresh media and then freezing it. Isolates can also taken from frozen populations after the experiment by thawing and streaking out the stock. | |||||||
Analyzing evolution experimentsThe end result of an evolution experiment is likely isolated, evolved clones that have acquired additional mutations relative to their ancestor. The goals, then, are usually to identify these mutations in the genome and determine what their functional consequences, if any, are. These can be accomplished by sequencing the isolates' genomes and comparing them to the ancestor with breseq and performing fitness assays or other functional assays (i.e. growth curve, minimum inhibitory concentration, biochemical assays, as appropriate), respectively. |
Evolution ExperimentsIntroductionAn evolution experiment consists of evolving one or more ancestor strains in laboratory culture and assessing the types and impacts of acquired mutations on the functions of that organism including its ability to reproduce, consume or produce compounds of interest, or resist particular stress conditions. The topics discussed on this page assume a general bacterial evolution experiment where a bacterial strain is serially passaged to fresh media over time. There are other types of evolution experiments including evolving phages, passaging pathogens and symbionts between hosts, and evolving multiple strains in co-culture that require additional considerations. In general there are two kinds of evolution experiments: adaptive evolution and mutation accumulation. In adaptive evolution experiments cells within a culture compete with each other and trend towards an increase in fitness. These experiments are useful for identifying mutations that improve a particular function, such as growth on a particular compound or resistance to a particular stress. Individuals with detrimental mutations are usually out-competed, thus the mutations evolved in these experiments is usually biased towards beneficial mutations. Mutation accumulation experiments, on the other hand, reduce or remove selection, allowing the persistence of all types of mutations and are thus useful for calculating overall mutation rates or examining the spectrum of possible mutations.Experimental designAn overall evolution experiment will likely involve three phases: 1) inoculating populations, 2) routine transfers, and 3) isolating clones and freezing stocks. 1) Inoculating populations Regardless of whether they are adaptive or mutation accumulation experiments, evolution experiments should be started by streaking the ancestral stock onto an agar plate and then initiating each population from a separate isolated colony. This reduces the impact of mutations that may already be present in the ancestral stock and ensures that the populations begin as single genotypes. After the first day of growth populations should be frozen so they can be sequenced and referred to later, in particular when determining whether identical mutations shared between samples evolved during the experiment or were present in the initial culture. 2) Routine transfers Over the course of an evolution experiment the populations will be transferred to fresh media to continue growing after their initial media has been used up. Designing an effective evolution experiment requires taking into account the impact of:
Analyzing evolution experiments | ||||||||
Changed: | ||||||||
< < | The end result of an evolution experiment is likely isolated, evolved clones that have acquired additional mutations relative to their ancestor. The goals, then, are usually to identify these mutations in the genome and determine what their functional consequences, if any, are. These can be accomplished by sequencing the isolates' genomes and comparing them to the ancestor with breseq and performing fitness assays or other functional assays, respectively. | |||||||
> > | The end result of an evolution experiment is likely isolated, evolved clones that have acquired additional mutations relative to their ancestor. The goals, then, are usually to identify these mutations in the genome and determine what their functional consequences, if any, are. These can be accomplished by sequencing the isolates' genomes and comparing them to the ancestor with breseq and performing fitness assays or other functional assays (i.e. growth curve, minimum inhibitory concentration, biochemical assays, as appropriate), respectively. |
Evolution ExperimentsIntroduction | ||||||||
Added: | ||||||||
> > | An evolution experiment consists of evolving one or more ancestor strains in laboratory culture and assessing the types and impacts of acquired mutations on the functions of that organism including its ability to reproduce, consume or produce compounds of interest, or resist particular stress conditions. The topics discussed on this page assume a general bacterial evolution experiment where a bacterial strain is serially passaged to fresh media over time. There are other types of evolution experiments including evolving phages, passaging pathogens and symbionts between hosts, and evolving multiple strains in co-culture that require additional considerations. | |||||||
In general there are two kinds of evolution experiments: adaptive evolution and mutation accumulation. In adaptive evolution experiments cells within a culture compete with each other and trend towards an increase in fitness. These experiments are useful for identifying mutations that improve a particular function, such as growth on a particular compound or resistance to a particular stress. Individuals with detrimental mutations are usually out-competed, thus the mutations evolved in these experiments is usually biased towards beneficial mutations. Mutation accumulation experiments, on the other hand, reduce or remove selection, allowing the persistence of all types of mutations and are thus useful for calculating overall mutation rates or examining the spectrum of possible mutations.
Experimental designAn overall evolution experiment will likely involve three phases: 1) inoculating populations, 2) routine transfers, and 3) isolating clones and freezing stocks. 1) Inoculating populations Regardless of whether they are adaptive or mutation accumulation experiments, evolution experiments should be started by streaking the ancestral stock onto an agar plate and then initiating each population from a separate isolated colony. This reduces the impact of mutations that may already be present in the ancestral stock and ensures that the populations begin as single genotypes. After the first day of growth populations should be frozen so they can be sequenced and referred to later, in particular when determining whether identical mutations shared between samples evolved during the experiment or were present in the initial culture. 2) Routine transfers Over the course of an evolution experiment the populations will be transferred to fresh media to continue growing after their initial media has been used up. Designing an effective evolution experiment requires taking into account the impact of:
Analyzing evolution experimentsThe end result of an evolution experiment is likely isolated, evolved clones that have acquired additional mutations relative to their ancestor. The goals, then, are usually to identify these mutations in the genome and determine what their functional consequences, if any, are. These can be accomplished by sequencing the isolates' genomes and comparing them to the ancestor with breseq and performing fitness assays or other functional assays, respectively. |
Evolution ExperimentsIntroductionIn general there are two kinds of evolution experiments: adaptive evolution and mutation accumulation. In adaptive evolution experiments cells within a culture compete with each other and trend towards an increase in fitness. These experiments are useful for identifying mutations that improve a particular function, such as growth on a particular compound or resistance to a particular stress. Individuals with detrimental mutations are usually out-competed, thus the mutations evolved in these experiments is usually biased towards beneficial mutations. Mutation accumulation experiments, on the other hand, reduce or remove selection, allowing the persistence of all types of mutations and are thus useful for calculating overall mutation rates or examining the spectrum of possible mutations.Experimental designAn overall evolution experiment will likely involve three phases: 1) inoculating populations, 2) routine transfers, and 3) isolating clones and freezing stocks. 1) Inoculating populations Regardless of whether they are adaptive or mutation accumulation experiments, evolution experiments should be started by streaking the ancestral stock onto an agar plate and then initiating each population from a separate isolated colony. This reduces the impact of mutations that may already be present in the ancestral stock and ensures that the populations begin as single genotypes. After the first day of growth populations should be frozen so they can be sequenced and referred to later, in particular when determining whether identical mutations shared between samples evolved during the experiment or were present in the initial culture. 2) Routine transfers Over the course of an evolution experiment the populations will be transferred to fresh media to continue growing after their initial media has been used up. Designing an effective evolution experiment requires taking into account the impact of:
Analyzing evolution experimentsThe end result of an evolution experiment is likely isolated, evolved clones that have acquired additional mutations relative to their ancestor. The goals, then, are usually to identify these mutations in the genome and determine what their functional consequences, if any, are. These can be accomplished by sequencing the isolates' genomes and comparing them to the ancestor with breseq and performing fitness assays or other functional assays, respectively. |