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Wheat Genetics Resource Center

Kansas State University
Department of Plant Pathology

4024 Throckmorton
1712 Claflin Road
Manhattan, KS 66506-5502
785-532-6176 (TEL)
785-532-5692 (FAX)
wgrc@k-state.edu

Wheat Genetics Resource Center
Kansas Wheat Innovation Center

1990 Kimball Ave
Manhattan, KS 66502
785-320-4383 (TEL)

 

HYBRIDIZATION PROCEDURE

Hybridization

Hybridization buffer [for 4 probes] 
ddH20
25.0 mL
20X SSPE
12.0 mL
100X Denhardt's solution
2.0 mL
20% SDS
1.0 mL

Salmon sperm DNA [10 mg/mL; boil for 5 minutes and set on ice 5 minutes before adding]

0.2 mL
dextran sulfate
2.0 g
TOTAL
40.2 mL
  1. Replace prehybridization buffer in each hybridization bottle with 10 mL of hybridization buffer.
  2. Add probe to hybridization bottle. NOTE: do not allow the probe to come in direct contact with the membrane when first adding the probe. Make sure the probe goes directly into the hybridization solution in the bottle.
  3. Allow probes to hybridize for 18–24 hours at 65°C.