Gentle hypochlorite treatment

Gentle hypochlorite treatment

1. Wash off plates of gravid hermaphrodites with 10-15 mls of M9 and put in 15 ml conical tube. Collect minimum of 4 or 5 plates in a tube.

2. Centrifuge @500rpm for about 3 minutes.

3. Remove spnt and resuspend animals in a solution of chlorox:M9 (3:7) - Use 1 ml total. Grind this solution in a glass mortar and pestle to disrupt adults (about 2.5-3 minutes or until whole adults are no longer visible).

4. Return animals to conical tube and wash out chlorox by adding 10mls of egg salts and centrifuging, @500rpm for about 5 minutes. Remove spnt and add about 1ml of a 1:7, chlorox:M9 solution to disrupt last remaining adults.

5. Wash embryos in M9 (10 mls), centrifuge @500 rpm for 5 mintues and then repeat wash with 10 mls of M9.

Regular hypochlorite treatment

1. Wash off plates of gravid hermaphrodites with 10-15 mls of water and put in 15 ml conical tube. Collect minimum of 4 or 5 plates in a tube.

2. Centrifuge @1700rpm for about 1 mintue.

3. Remove spnt and resuspend animals in 15ml [10% chlorox, 1 N NaOH]. Let incubate at room temperature for 3 minutes with occasional shaking.

4. Pellet worms by centrifuging @1700rpm for about 1 minute. Remove spnt and add another 15ml [10% chlorox, 1 N NaOH]. Let incubate at room temperature for 3 minutes with occasional shaking.

5. Wash embryos in M9 (15 mls), centrifuge @1700rpm for about 1 mintue, and repeat wash with 15 mls of M9.