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Plasmid Preparation column is blocked

Plasmid Preparation column is blocked - Molecular Cloning Forum

Plasmid Preparation column is blocked - DNA cloning forum. Discuss the cloning of plasmids, vectors, DNA, RNA, cDNA, proteins, and other molecules for expression, sub-cloning, and other methods. Also discuss digestion, ligation, transformation, plasmid preparation, cDNA library screening, and competent cells here.


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Old 05-11-2011, 05:47 PM
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Exclamation Plasmid Preparation column is blocked



Hi everybody,

at the first time i use the maxi kit from this company i could get the dna plasmid. Today i did exactly the same but my column got blocked. The lysat liquid after soltion III flows very slowly. I need 4 hours until it flows through completely. Can i still use the dna afterward if i finally done the maxi prep? I plan for the next time after the centrifugation to transfer the lysat into a filter paper and let it filtered and look whether precipitates are still there. Until i get precipitate free lysat. Then i apply to the maxi column for binding the plasmid dna. Do you have further suggestions? I would be very happy if you do

Smiley
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Old 05-19-2011, 06:55 PM
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Default Re: Plasmid Preparation column is blocked

It's possible that you sheared the DNA during the lysis phase. After adding the neutralization buffer, gently invert/roll the bottle, then centrifuge.
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Old 05-20-2011, 06:19 AM
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Default Re: Plasmid Preparation column is blocked

Hola, we use a gauze in the top of colum to retain any solid of DNA suspension and some DNA purification kits (giga? quiagen) has filter paper as you suggest to clarify the DNA solution after centrifugation. Buena suerte
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Old 06-18-2012, 06:43 AM
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Default Re: Plasmid Preparation column is blocked

thanks your help
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