9楼: Originally posted by wxl.1989822 at 2014-03-06 22:04:23
构建成了 酶切位点估计没有问题...

6楼: Originally posted by 西门吹雪170 at 2014-03-06 21:42:46
这也是经常遇到的问题，重组质粒大小正确，进行PCR验证也有出现目的条带，但是就是酶切不出条带，建议你测一下重组质粒的浓度，然后按最终反应体系质粒的浓度约为20ug/uL进行计算所需加样的量，试试看吧，一般这样是 ...

12楼: Originally posted by wxl.1989822 at 2014-03-07 09:16:39
20ug/ul?这么大...

14楼: Originally posted by 鬼羽帝魂 at 2014-03-07 10:54:39
你每次酶切的重组质粒大约有多少ng？

13楼: Originally posted by 西门吹雪170 at 2014-03-07 09:37:30
这还大啊，我们同学有时用的比这个浓度还大，都切开了...

15楼: Originally posted by wxl.1989822 at 2014-03-07 15:14:50
我测了下浓度大概在60ng/ul,我加的有16 14 12 8 4ul 等，在20ul 的酶切体系中。...

18楼: Originally posted by 鬼羽帝魂 at 2014-03-07 17:06:42
试下8ul，切6h。...

19楼: Originally posted by wxl.1989822 at 2014-03-07 18:28:42
切这么久 我觉得应该是我的质粒浓度太低 酶活性过高 切碎了...