点赞

收藏

分享

E.Z.N.A. Cycle-Pure Kit Spin Protocol

互联网2013-11-13

2553

实验试剂

1. Optional: Sterile deionized water

2. Absolute ethanol (~ 96-100%)

实验设备

1. Microcentrifuge capable of at least 13,000x g

2. Nuclease-free 1.5ml centrifuge tubes

实验步骤

1. Perform agarose gel/ethidium bromide electrophoresis to analyze PCR product.

2. Determine the volume of the PCR reaction, transfer the sample into a clean 1.5ml microcentrifuge tube, and add 4-5 volumes of Buffer CP. For PCR products < 200bp add 6 volumes of Buffer CP.

3. Vortex thoroughly to mix. Briefly spin the tube to collect any drops from the inside of the lid.

4. Place a HiBind® DNA column in a provided 2ml collection tube. Add 100ul Equilibration Buffer to the column. Incubate at room temperature for 4 minutes. Spin at maximum speed for 20 seconds.

5. Apply the sample to the HiBind® DNA column and centrifuge at 10,000x g for 1 min at room temperature. Discard the flow-through.

6. Wash the HiBind® DNA column by adding 700ul of DNA Wash Buffer diluted with absolute ethanol and centrifuge as above.

IMPORTANT: DNA Wash Buffer must be diluted with absolute ethanol before use. Refer to label for instructions. If refrigerated, DNA Wash Buffer must be brought to room temperature before use.

7. Discard liquid and repeat Step 7 using 500ul of DNA Wash Buffer.

8. Discard liquid and centrifuge the empty HiBind® DNA column for 2 min at maxi speed (13,000x g) to dry the column matrix. This is critical for good yields.

9. Place HiBind® DNA column into a clean 1.5ml microcentrifuge tube. Add 30-50ul (depending on desired concentration of final product) of Elution Buffer (10mM Tris, pH8.5) or water directly onto the column matrix and centrifuge for 1 min at 13,000x g to elute DNA.This represents approximately 80-90% of bound DNA. An optional second elution will yield any residual DNA, though at a lower concentration.

10. Yield and quality of DNA: Determine the absorbance of an appropriate dilution of the sample at 260nm and then at 280nm. The DNA concentration is calculated as follows:

260 DNA concentration = A x 50 x (Dilution Factor) ug/ml

Fragments greater than 500bp in length can routinely be purified at > 80% yield. 260 Bands ranging from 100bp to 500bp gives yields of 60%- 90%. The ratio of (A )/280 (A ) is an indication of nucleic acid purity. Alternatively, yield (as well as quality) can sometimes be best determined by agarose gel/ethidium bromide electrophoresis.

相关产品推荐

RNAprep Pure多糖多酚植物总RNA提取试剂盒(DP441)

RNAprep Pure多糖多酚植物总RNA提取试剂盒(DP441)

￥1480

SPIN1/SPIN1蛋白Recombinant Human Spindlin-1 (SPIN1)重组蛋白Ovarian cancer-related protein Spindlin1蛋白

SPIN1/SPIN1蛋白Recombinant Human Spindlin-1 (SPIN1)重组蛋白Ovarian cancer-related protein Spindlin1蛋白

￥1344

KTO: KIT A643 VALIDATION PROTOCOL (L)-Anatel A643a Accessories

KTO: KIT A643 VALIDATION PROTOCOL (L)-Anatel A643a Accessories

￥50094

糖组学分析| 糖蛋白质组学分析服务| 蛋白质N-糖基化分析| 蛋白质O-糖基化分析

糖组学分析| 糖蛋白质组学分析服务| 蛋白质N-糖基化分析| 蛋白质O-糖基化分析

询价

Mouse IFN-γ High Sensitivity ELISA Kit（小鼠干扰素IFN-γ高敏试剂盒）

Mouse IFN-γ High Sensitivity ELISA Kit（小鼠干扰素IFN-γ高敏试剂盒）

￥1600

相关问答

问

pierce albumin depletion kit

问

WB显出膜没有条带

问

2小时能完成Western Blot实验，可能吗？

相关方法

利用 Ambion 的 MicroPoly(A)Pure 试剂盒分离 mRNA 实验

2024-05-14

质粒 D N A 的制备

2024-05-14

使用 HC ExpressArray Kit 进行杂交与检测实验

2024-05-14

推荐阅读

E.Z.N.A.® Endo-free Plasmid Mini Kit I Spin Protocol

E.Z.N.A. Endo-Free Plasmid Mini Kit II Spin Protocol

E.Z.N.A.® Viral RNA Spin Protocol

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