Difference between revisions of "Performing Drosophila Crosses"
From Bridges Lab Protocols
| Line 1: | Line 1: | ||
| + | ==Preparing the Crosses== | ||
*Collect virgin females with appropriate marker and put into a new vial. | *Collect virgin females with appropriate marker and put into a new vial. | ||
*Store the vials from which virgins are collected at 18° C when not using. | *Store the vials from which virgins are collected at 18° C when not using. | ||
| Line 4: | Line 5: | ||
*Once 10 virgin females from a strain have been collected, add 4 males with the appropriate marker from a different strain. | *Once 10 virgin females from a strain have been collected, add 4 males with the appropriate marker from a different strain. | ||
*Label the vial to indicate the cross type and generation (A1 = the first generation of the first cross). | *Label the vial to indicate the cross type and generation (A1 = the first generation of the first cross). | ||
| − | *Store the crosses in an incubator. | + | *Store the crosses in an appropriate temperature-controlled incubator. |
*After 5 days, flip the 14 flies into a new vial (there should be eggs visible at the surface of the food). | *After 5 days, flip the 14 flies into a new vial (there should be eggs visible at the surface of the food). | ||
| − | *Label the vial appropriately (A2 = the second generation of the first cross). | + | *Label the new vial appropriately (A2 = the second generation of the first cross). |
| − | *Repeat this cycle until enough progeny from the crosses have been produced | + | *Repeat this cycle until enough progeny from the crosses have been produced. |
| − | * The progeny will start eclosing after ~10 days. | + | |
| + | ==Collecting the Progeny== | ||
| + | * The progeny from the crosses will start eclosing after ~10 days. | ||
* Record the date that the flies begin eclosing. | * Record the date that the flies begin eclosing. | ||
* Once there are at least 20 flies in the vial, collect the progeny and sort according to phenotype. | * Once there are at least 20 flies in the vial, collect the progeny and sort according to phenotype. | ||
| Line 14: | Line 17: | ||
* Place the males and females into separate vials. | * Place the males and females into separate vials. | ||
* There should be 5 - 15 progeny flies in each vial. | * There should be 5 - 15 progeny flies in each vial. | ||
| − | **If necessary, separate/combine phenotypes to have an appropriate number of flies in the vial. | + | **If necessary, separate/combine phenotypes (but not gender) to have an appropriate number of flies in the vial. |
* Label the vial with: | * Label the vial with: | ||
#Cross type/generation | #Cross type/generation | ||
| Line 22: | Line 25: | ||
#date born | #date born | ||
#date added to vial. | #date added to vial. | ||
| − | * Keep the progeny vials in an incubator. | + | * Keep the progeny vials in an appropriate incubator. |
| − | * Repeat this process during the six days following the date of first eclosure | + | * Repeat this process during the six days following the date of first eclosure. |
| − | **Wait to sort until there are at least 20 flies in the vial (usually 2 to 3 times per vial) | + | **Wait to sort until there are at least 20 flies in the vial (usually 2 to 3 times per vial). |
| − | *After six days, toss the vial | + | **Record the most likely date of birth before each sorting. |
| − | * Flip all progeny vials twice a week | + | *After six days, toss the vial. |
| + | |||
| + | ==Recording Deaths of the Progeny== | ||
| + | * Flip all progeny vials into fresh vials twice a week (every 3-4 days). | ||
| + | *Record information on any dead flies found while flipping: | ||
| + | #Cross type | ||
| + | #Gender | ||
| + | #Phenotype | ||
| + | #Date born | ||
| + | #Date of death | ||
Latest revision as of 17:35, 20 June 2013
Preparing the Crosses
- Collect virgin females with appropriate marker and put into a new vial.
- Store the vials from which virgins are collected at 18° C when not using.
- Store vials from which males are collected at room temperature.
- Once 10 virgin females from a strain have been collected, add 4 males with the appropriate marker from a different strain.
- Label the vial to indicate the cross type and generation (A1 = the first generation of the first cross).
- Store the crosses in an appropriate temperature-controlled incubator.
- After 5 days, flip the 14 flies into a new vial (there should be eggs visible at the surface of the food).
- Label the new vial appropriately (A2 = the second generation of the first cross).
- Repeat this cycle until enough progeny from the crosses have been produced.
Collecting the Progeny
- The progeny from the crosses will start eclosing after ~10 days.
- Record the date that the flies begin eclosing.
- Once there are at least 20 flies in the vial, collect the progeny and sort according to phenotype.
- Record the number of flies of each phenotype born.
- Place the males and females into separate vials.
- There should be 5 - 15 progeny flies in each vial.
- If necessary, separate/combine phenotypes (but not gender) to have an appropriate number of flies in the vial.
- Label the vial with:
- Cross type/generation
- gender
- number of flies
- phenotype
- date born
- date added to vial.
- Keep the progeny vials in an appropriate incubator.
- Repeat this process during the six days following the date of first eclosure.
- Wait to sort until there are at least 20 flies in the vial (usually 2 to 3 times per vial).
- Record the most likely date of birth before each sorting.
- After six days, toss the vial.
Recording Deaths of the Progeny
- Flip all progeny vials into fresh vials twice a week (every 3-4 days).
- Record information on any dead flies found while flipping:
- Cross type
- Gender
- Phenotype
- Date born
- Date of death
