How to know the exact plasmid size?
Posted 27 March 2009 - 06:31 PM
i m now working onto some new isolated bacteria on my own and i have no idea that wat kind of plasmid in inside them.
i have extracted out the plasmid DNA and did a gel run.
and it show me with some multiple bands, yes, i know it's common for plasmid DNA due to its conformation (supercoiled, nicked and linear), so it gives me different migration pattern.
my question is,
how do i determine which is the EXACT size of the plasmid that i extracted ? since it migrates differently though at same size ?
pls correct me if i am wrong.
Posted 27 March 2009 - 06:41 PM
Posted 27 March 2009 - 09:25 PM
Posted 27 March 2009 - 10:19 PM
Run standard vectors without digesting and compare their mobility and estimate aproximate size of yr plasmid.
Hope it helps.
Posted 27 March 2009 - 11:35 PM
but my question is that if your bacteria strain is already known, then can't you find that plasmid sequence at Genebank or somewhere to figure out what RE would give you only 1 linear band?
but different bacteria may possess different plasmid one, right ? then how could i be so sure that which plasmid that it have ?
Posted 28 March 2009 - 02:56 AM
You might get several DNA fragments from the digest. Just sum the sizes of the DNA fragments.
Repeat this for several different enzymes just in case an enzyme does not cut or cuts too frequently.
Posted 28 March 2009 - 03:10 AM