Slide Spotting and SYBR Staining

The Amersham Biosciences Lucidea Array Spotter (see Note 5) is used to deposit DNA on CodeLink Activated Slides (Fig. 6). First the slides are washed and blocked against nonspecific binding and then processed for staining.

1. Condition the pen set by washing several times (with standard recommended buffers) before starting the spotting run.

2. Upon completion of spotting, slides should be left in the machine at 70% humidity for at least 30 min.

3. Place the slides in an open rack in a tightly sealed saturated NaCl humidification chamber overnight. Slides are then ready for use.

4. Wash with Blocking solution for 30 min. Note: SDS is not added.

5. Wash with postcoupling wash solution.

6. Pipet 200 pL of a 1:2500 dilution of SYBR Green I or SYBR Gold in 4X SSC onto slide spotting area. Cover with Parafilm and incubate in a humid chamber for 45 min.

7. Wash with 4X SSC/0.1% Triton X-100 for 5 min twice.

8. Wash with 4X SSC for 5 min twice.

9. Dry slides by centrifugation at 200g for 3 min.

10. Scan in Axon GenePix 4000B scanner (see Subheading 3.9.).

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