关于DNA LADDR提取柱子的选择

问： 对比了DNA ladder的实验方法和一些公司的dna ladder提取试剂盒，发现关键的区别就在回收DNA LADDER的柱子上。由于经费有限，请问提取质粒或是回收试剂盒中的柱子能够用于回收DNA LADDER吗？

答：

实验操作步骤：

1、 Harvest cells and pre-cold PBS rinsing once, 500 g centrifuge 5 min Room Temprature

2、 Pellet resuspended in 250 μl TE buffer containing 1μg/μl RNase A

3、 Cells were lysed by adding an equal volume of 1.2% SDS and gentle mixing by inversion(6-8 times)

4、 Incubate 5 min, 350 μl precipitation solution were added, mixed 8-10 times and put on ice for 15 min

5、 Spin 14000g for 15 min Room Temperature and clear supernatant (700 μl) transfer to miniprep column

6、 Spin at 14000 g for 1 min, washed with 700 μl wash buffer once

7、 Spin empty column to discard the rest ethonal

8、 30-50 μl TE(pH8.0) elute the DNA fragments

9、 1.6% agarose gel separated at 30V for 10min, and 80 V for 50 min

10、 EB staining and photographed

实验材料与试剂配制：

1、 TE buffer

2、 CsCl containing precipitation buffer:3M CsCl,1M Potassium acetate,0.67M acetic acid

3、 Wash buffer:80mM potassium acetate,10mM Tris-Cl,pH7.5,40uM EDTA,60% ethanol

4、 1.6% Agarose gel

5、 Miniprep column (QIAprep Spin Miniprep Kit, QIAGEN)

6、 ethidium bromide (10 μl/100ml)

我们实验室是这样做的，如果药物或者处理能够诱导凋亡，都还做得蛮好的，供参考。