Performing Drosophila 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 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 vial appropriately (A2 = the second generation of the first cross).
• Repeat this cycle until enough progeny from the crosses have been produced or until there are no parent males left.
• The progeny 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 to have an appropriate number of flies in the vial.
• Label the vial with:

1. Cross type/generation
2. gender
3. number of flies
4. phenotype
5. date born
6. date added to vial.

• Keep the progeny vials in an 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)
• After six days, toss the vial
• Flip all progeny vials twice a week