• 我要登录|
  • 免费注册
    |
  • 我的丁香通
    • 企业机构:
    • 成为企业机构
    • 个人用户:
    • 个人中心
  • 移动端
    移动端
丁香通 logo丁香实验_LOGO
搜实验

    大家都在搜

      大家都在搜

        0 人通过求购买到了急需的产品
        免费发布求购
        发布求购
        点赞
        收藏
        wx-share
        分享

        Silver Staining Protocol for Acrylamide Gels

        互联网

        651
        相关专题
         

         

        Gel Mix

         

         

        For 1 litre solution

         

         

        Urea (AG)

        453 gms

         

         

        10X TBE

        50 ml

         

         

        40%(19:1 ratio) Acrylamide : bisacrylamide

        125 ml

         

         

        Distilled water

        (make up to 1 litre)

         

        Electrophoresis

        The small and big glass plates wiped thoroughly with detergent and acetone and allowed it to dry

        Repel silane is applied to the big glass plate and bind silane is applied to the small glass plate uniformly.

        The spacer (0.4 mm) is kept at the edge of the big glass on the repellent applied side and silane applied side of small glass plate is placed over it.

        Take care to align both the glass plates correctly

        Take 60 ml of gel solution, 250ul (10mg/ml) ammonium persulfate and 80 l TEMED and mix briefly

        Pour the gel between the glass plates and allow it to settle for a minimum of 1 hr.

        Fix the gel plates on to the electrophoretic unit and clean the urea with buffer.

        Prick the gel with 62 well combs.

        Pre warm the gel at 40W for 20 min.

        Then clean the residual urea from the wells by taking 50 ml 0.5 X buffer in a syringe.

        Add equal volume of dye solution to the PCR samples and mix thoroughly.

        DeNature the PCR products at 94ºC for 3 min just before loading and immediately keep it in ice.

        Load 4-5 µl of PCR products in each well.

        The running conditions are 2 hrs at 40 constant Watts.

        After gel running is completed, the combs are separated and the big glass plate is removed. The gel sticking onto the small glass plate will be used for silver staining

        Silver Staining for DNA visualization

        Gently place the gel in 10% (v/v) glacial acetic acid for 30 min at room temp.

        Rinse the gel in deionized water twice for about 2 min each.

        Immerse the gel in silver staining solution (250 mg silver nitrate and 375 µL formaldehyde in 250 ml water) for 20 min.

        Pour out the silver stain solution and wash the gel quickly with deionized water with in 10 seconds.

        Immerse the gel in an ice-cold developer solution (10oC)

        ad image
        提问
        扫一扫
        丁香实验小程序二维码
        实验小助手
        丁香实验公众号二维码
        扫码领资料
        反馈
        TOP
        打开小程序