DNA Hybridization - Church Method

Jeff Lawrence

1. To prepare Hybridization Solution, mix in order:

	13 mL	ddH₂O
	2 mL	500 mM EDTA
	1 g	Grade V BSA
	50 mL	1 M NaHPO4, pH 7.2
	35 mL	20% SDS

2. Prehybridize filters at 65° for 2 hr. Remove solution and replace with hybridization solution and labeled probe. Incubate at 65° for 12 hr.

3. Remove filter and place in 100 mL of:

	2X	SSC	100 mL	20X SSC
	1 %	SDS	100 mL	10 % SDS
			800 mL	ddH₂O

4. Rinse at 22C for 15 min. Repeat.

5. If filter is uniformly radioactive, rinse at 60° for 30 min in:

	0.1 X	SSC
	1 %	SDS

6. Expose to X-Ray film.

Last Update: Thursday June 19 2014

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Eric Kofoid eckofoid at ucdavis.edu