Protocol Online logo
Top : Forum Archives: : Molecular Cloning

What way would you suggest to clean PCR products? - (Oct/06/2005 )

Hi,I am planning to do PCR soon.However, I do not know the correct and efficient way to clean my PCR products. After PCR, I will continue with cutting the PCR products with restriction enzyme. My template for PCR is the NP gene from NDV. Any suggestion to clean the PCR products?


We use a kit -- the QIAquick PCR Purification Kit. It works well.

Just as an aside, be sure to gel-purify your PCR product after you digest it...


Sometimes it's ok to digest ur PCR product without purification, depending on ur PCR mix and the RE enzymes that u r using
Run a trial to check with this, if it's ok without purification step, u can save quite some money there wink.gif


Thanks for the assistance.
By the way,Homebrew did mentioned about gel-purifying the PCR products after digestion,what does gel-purifying mean?Is it about performing agarose gel electrophoresis to get my desire PCR products and get rid of other contaminants?


QUOTE (natural_leaves @ Oct 6 2005, 09:35 PM)
what does gel-purifying mean?

Coincidently, I'm still at the lab here at 8:45 PM doing exactly this...biggrin.gif

I'm going to clone two genes. I did two PCRs overnight last night (50 ul reactions). This morning, I ran a small sample (8 ul) out on a gel, and saw that the PCRs worked well. I took both the PCRs and cleaned them up with the above mentioned PCR clean up kit, digested the products with Bam HI (Bam HI sites were engineered into the 5' ends of my primers), and loaded the whole restriction digest on another agarose gel. I ran that gel out (this is the gel purification part), and excised the bands. I used Qiagen's gel extraction kit to recover the DNA, and I'm setting up my ligations now to go overnight...