Is there any way to remove chromosomal DNA from plasmid DNA? - (Sep/23/2008 )
Hello,
I isolate plasmid DNA from E.COLI by phenol-chloroform extraction, alkaline lysis. After running gel, I saw some smear(big band) around 20kb. Is it genomic DNA? Is there any way to remove those chromosomal DNA? Thank you!
it is quite likely genomic DNA, although it could also be protein junk if the prep isn't very clean.
It depend on how much DNA. From the entire midiprep, probably not in cost/time efficient way.
If it is for a fragment to be clone, probably yes. One could use gel extraction.
I isolate plasmid DNA from E.COLI by phenol-chloroform extraction, alkaline lysis. After running gel, I saw some smear(big band) around 20kb. Is it genomic DNA? Is there any way to remove those chromosomal DNA? Thank you!
What the other guy said: do a gelextraction of the entire plasmid (if that's what you need). You'll find it on top of the gel
I isolate plasmid DNA from E.COLI by phenol-chloroform extraction, alkaline lysis. After running gel, I saw some smear(big band) around 20kb. Is it genomic DNA? Is there any way to remove those chromosomal DNA? Thank you!
What the other guy said: do a gelextraction of the entire plasmid (if that's what you need). You'll find it on top of the gel
Hey while doing extraction, i see an extra band around 5 kb, i dont know where it comes from? I have done the extraction with phenol-chloroform alkaline lysis six times and each time i get that extra band. My friends say this is some sort of contamination. Can anyone help?
What is the size of your plasmid? Could that extra band be supercoiled or nicked DNA.
Its about 4kb. I dont know should i use that that plasmid or not.
One more thing when i was making RNA with mmessage mmachine kit, and when i ran the gel, i did not
get that RNA band but rather obtanied some bands about the size of my plasmid DNA, i am not sur e what actually it is?
My collegues also dont know about that, they say its mystery but i want the convincingg answer for this.
Can anyone help????????
try linearizing the plasmid DNA with restriction enzyme that's known to cut it only once and see if that mysterious 20kb band is gone or nt.