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Molecular Cloning Forum 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.


Alkali lysis and transformation

Alkali lysis and transformation - Molecular Cloning Forum

Alkali lysis and transformation - 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 10-04-2010, 12:58 AM
Pipette Filler
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Default Alkali lysis and transformation



Hi,
I was wondering why the alkali lysis plasmid isolation method is effective in producing a cleared lysate?
Also, i was wondering if the reason for a triple plasmid transformation having a lower colony count compared to a single or double transformation is, just basically because the membrane is somewhat congested by all the plasmids being present and the larger plasmids would be blocking the smaller plasmids form getting through the pores.
Thanks
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Old 11-16-2010, 08:58 PM
Pipette Filler
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Default Re: Alkali lysis and transformation

Hi

Transformation of cells all depends upon the chance. If I where on your position. I would do the double transformation first. Later screen the cells for plasmid, then go for making them competent again with contain previous plasmid then I will do the third transformation using different antibiotic marker

Tedious but effective
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