IGEM:Peking/2007/Switch: DNA ligation

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Revision as of 05:29, 18 October 2007 by Chen Chongyi (talk | contribs) (New page: ==连接== 1.普通连接的体系为10uL,其中片断+载体5uL,Ligation Kit 5uL。 2.Ligation Kit需在冰盒中溶化。分装至1.5mL离心管中使用。 3.先加片断...)
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连接

1.普通连接的体系为10uL,其中片断+载体5uL,Ligation Kit 5uL。

2.Ligation Kit需在冰盒中溶化。分装至1.5mL离心管中使用。

3.先加片断,后加载体。

4.管口用Parafilm封好。

5.16℃反应1hr以上。

注意:连接酶25℃达到最大活性,3min失活。4℃活性较低,但可以保持过夜。16℃的性质在二者之间。4℃连接过夜的效果最好。根据片断与载体连接的难易程度选择连接的温度与时间。