I have three samples. One is a product obtained from CO1 primer with band of interest at around 350-380 bp, and two products obtained from ITS 1 & 2 primers with bands of interest at 340-375 bp.
I wish to perform gel extraction/elution of DNA for all three and send it for sanger sequencing (ABI 3730)
Kindly suggest what percentage of agarose I should use and how thick I should pour the gel to get better resolved bands and good eluted DNA for sanger sequencing?
Hi Anuradha Joshi, considering the PCR product size you should prepare a gel of 2-2.5%. The thickness of the gel depends on the size of your wells because you need to load as much as PCR products. From my experience, if you load around 10ug of the sample into the gel, your extraction yield will be around 1-2ug. Use the QIAquick Gel Extraction Kit (Cat no.28704) and make sure the final elution volume is 10-12uL. Also, make sure you dissolve the gel completely in the first step.
Best!
-Pushkal
@Pushkal Singuvani Ramesh
Thank you for your answer!
By 10ug of sample do you mean the final DNA concentration after amplification?
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