Preparation of G+A Marker Procedure 1、Add the following to a sterile microcentrifuge tube: 2、Add 1ul of 4% Formic Acid and incubate for 25 min at 37℃ . 3、During this incubation, add 15 ul of stock piperidine to 135 ul of water to prepare a 1M piperidine solution. 4、Place the tube containing the formic acid reaction on ice, add all 150 ul of the diluted (1M) piperidine solution and incubate for 30 min at 90℃. 5、Place this reaction on ice for 5min, add 1ml of n-butanol and vortex. 6、Centrifuge for 2mi8n at high speed to pellet the DNA. Remove the supernatant and add 150ul of 1% SDS to the pellet. 7、Add 1ml of n-butanol, vortex vigorously and centrifuge at high speed for 2 min. Carefully remove the supernatant. 8、Add 0.5 ml of n-butanol to rinse the pellet, and carefully remove the supernatant. Repeat this rinse step once. 9、Dry the pellet under vacuum for 10min, adds 5-10ul of loading dye, and mix well. Place at 20℃ until required. This sample may be stored up to two weeks at 20℃. This protocol was adopted from Amersham footprinting kit instruction |
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