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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.|
|View Poll Results: plasmid isolation problem|
|Is the colonies of dh5 alpha appaered as pink?||0||0%|
|is there any different protocol for plasmid isolation from dh5 alpha.||0||0%|
|transformation efficiency of xl1 blue cells.||0||0%|
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plasmid isolation problem
I am facing one problem with plasmid isolation. nw i am doing one project wrk on pcr mutagnesis.
transformation of the pcr product gave one colony after 12 hrs. i isolated the dna from the colony( XL-1 blue supercompetent cells were used for transformation)
But the concentration is very less, i.e; appro.. 25ng/ul
so i again transform the isolated plasmid to DH5 alpha cells to get more.
But it also gives lowest concention.I have done these things repeatedly.
I am using the phenol chloroform method for the plasmid isolation.According to the competent cells used,is there any change in the protocol..
please help me, if i didn't get enough plasmid, i am not able to send it for sequencing.
|isolation , plasmid , problem|
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