Let's say, I amplified a gene from a unknown bacteria and want to determine whether this gene located in plasmids or chromosome.
Dear Daniel, While I agree with the suggestions given above, you might be minded of the sort of techniques used in the 1980'2 and 1990's to look at plasmid-borne genes - it is not too hard to run lysed-cell samples on a gel to reasonably separate plasmid and genomic DNA, and then to either perform a Southern transfer and blot with the plasmid gene, or to isolate a small sample of plasmid DNA for PCR analysis. Whilst all of the modern techniques work - and provide in some cases a lot of additional information - the simple old-fashioned techniques are generally faster and allow you to quickly test a hypothesis without committing significant resources. Regards, Andrew.
Extract plasmids and do Northern.
Or flaking PCR and mapping to chromosome.
I agree with Juan Manuel. You can do a digestion with a plasmid safe ATP dependant DNAse. After that you can run the digestion on an agarose gel to isolate the plasmid and proceed to PCR, southern or sequencing.
Dear Daniel
I would like to say, after these methods which are said above, to assure about preventing the contamination of bacterial DNA, you can use transformation technique in order to transform the plasmid in different species or even genus which you are sure they haven't that gene.
after preparing the pure cell culture of that bacterium, you can use PCR technique to determine if that gene located in the plasmid or not.
For most of the mentioned techniques you need to culture your strain. If you are able to obtain a pure culture of your unknown bacterial strain you can easily perform whole genome sequencing (WGS). This will answer your question as well as many questions more. WGS for a bacterial strain can be performed for about $200.
Dear Daniel, While I agree with the suggestions given above, you might be minded of the sort of techniques used in the 1980'2 and 1990's to look at plasmid-borne genes - it is not too hard to run lysed-cell samples on a gel to reasonably separate plasmid and genomic DNA, and then to either perform a Southern transfer and blot with the plasmid gene, or to isolate a small sample of plasmid DNA for PCR analysis. Whilst all of the modern techniques work - and provide in some cases a lot of additional information - the simple old-fashioned techniques are generally faster and allow you to quickly test a hypothesis without committing significant resources. Regards, Andrew.
There are many suggestions and answers already there, I would like to add one simple fact that determine the copy number of the gene (may be real time PCR), if its more than two, it has to be plasmid origin.
Extract the plasmid/genomic DNA. Isolate on the agarose gel then do PCR or southern blotting
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