Materials and Reagents

1. 15 ml Falcon tubes (Thermo Fisher Scientific, Thermo Scientific^TM, catalog number: 339651 )
   
2. 50 ml Falcon tubes (Thermo Fisher Scientific, Thermo Scientific^TM, catalog number: 339653 )
   
3. 225 ml Falcon bottles (Corning, Falcon^®, catalog number: 352075 )
   
4. Ascaris
   Ascaris can be collected from slaughterhouses that process several thousand pigs a day and use the small intestines to make sausage casings. Ascaris can be picked up by hands (with gloves) from contents of intestine, which are pushed out to a tank by machines. Usually, it takes two persons 3-5 h to collect ~1,000 worms. Both fresh tissues and embryos can be obtained from these live worms. Female Ascaris can also be ordered from Carolina Biological, Living Zoology Department. These females are shipped dead on ice and the Ascaris zygotes can be obtained from the proximal uteri
   
5. Sodium hydroxide (NaOH) (Fisher Scientific, catalog number: S318-1 )
   
6. H₂O (purified by Milli-Q^® Integral Water Purification System)
   
7. DAPI (Santa Cruz Biotechnology, catalog number: sc-3598 )
   
8. Liquid nitrogen (Airgas, catalog number: NI NF160LT22 )
   
9. Phenylmethanesulfonyl fluoride (PMSF) (Thermo Fisher Scientific, Thermo Scientific^TM, catalog number: 36978 )
   
10. cOmplete^TM protease inhibitor cocktail (Roche Diagnostics, catalog number: 0493116001 )
    
11. Sodium chloride (NaCl) (Fisher Scientific, catalog number: S271-3 )
    
12. Potassium chloride (KCl) (Fisher Scientific, catalog number: P217-500 )
    
13. Sodium phosphate dibasic (Na₂HPO₄) (Fisher Scientific, catalog number: BP350-500 )
    
14. Potassium phosphate monobasic (KH₂PO₄) (Fisher Scientific, catalog number: P285-500 )
    
15. Hydrochloric acid (HCl)
    
16. 1 M Tris-HCl, pH 8.0 (Thermo Fisher Scientific, Ambion^TM, catalog number: AM9856 )
    
17. 1 M Tris-HCl, pH 7.0 (Thermo Fisher Scientific, Ambion^TM, catalog number: AM9851 )
    
18. Sodium hypochlorite solution (Fisher Scientific, catalog number: SS290-1 )
    
19. Potassium hydroxide (KOH) (Fisher Scientific, catalog number: P250-1 )
    
20. Magnesium chloride (MgCl₂) (Fisher Scientific, catalog number: M33-500 )
    
21. EGTA (Thermo Fisher Scientific, USB, catalog number: 15703 )
    
22. Sucrose (Fisher Scientific, catalog number: BP220-212 )
    
23. DTT (Fisher Scientific, catalog number: BP172-5 )
    
24. Spermine (Sigma-Aldrich, catalog number: S3256 )
    
25. Spermidine (Sigma-Aldrich, catalog number: S2626 )
    
26. Nonidet P-40 (NP-40) (Alfa Aesar, Affymetrix/USB, catalog number: J19628 )
    
27. Triton X-100 (Promega, catalog number: H5142 )
    
28. Glycerol (Fisher Scientific, catalog number: BP229-1 )
    
29. 2-mercaptoethanol (Sigma-Aldrich, catalog number: M6250-10ML )
    
30. EDTA (Thermo Fisher Scientific, Thermo Scientific^TM, catalog number: 17892 )
    
31. PBS buffer (see Recipes)
    
32. 1 M Tris-HCl, pH 7.8 (see Recipes)
    
33. KOH/Bleach (0.4 N KOH/1.4% sodium hypochlorite) (see Recipes)
    
34. Nuclei extraction buffer A (see Recipes)
    
35. Nuclei extraction buffer B (see Recipes)
    
36. Nuclei extraction buffer C (see Recipes)
    
37. Nuclei storage buffer (see Recipes)
    

Equipment

1. Erlenmeyer flask (Thermo Fisher Scientific, Thermo Scientific^TM, catalog number: 4112-0125 , 4112-0250 or 4112-0500 )
   
2. 1 L beaker (Thermo Fisher Scientific, Thermo Scientific^TM, catalog number: 12011000 )
   
3. 4 L plastic beaker (Thermo Fisher Scientific, Thermo Scientific^TM, catalog number: 12014000 )
   
4. Shaking incubator (example, Lab-Line, model: 4629 )
   
5. Fluorescence microscope (example, Nikon Instruments, model: TS100 )
   
6. Refrigerated centrifuge (example, Thermo Fisher Scientific, model: Sorvall^TM Legend^TM XTR )
   
7. Metal dounce homogenizer (WHEATON, catalog number: 357574 )
   
8. Mortar and pestle (example, Fisher Scientific, catalog numbers: S02591 and S02594 )
   

Procedure