DNA agarose gel: one than one chromosome, one band.

  • Thread starter Thread starter nobahar
  • Start date Start date
  • Tags Tags
    Band Dna
Click For Summary
SUMMARY

The discussion centers on the observation of a single smeared band in an agarose gel electrophoresis of undigested DNA from an organism with multiple chromosomes. This phenomenon occurs because larger DNA molecules, particularly those over 15-20kb, migrate together in a size-independent manner due to entanglement. The user, Nobahar, highlights that while this explains the single band for multiple chromosomes, the presence of insertions can affect migration rates, suggesting that entanglement allows for additive effects in migration. The conversation concludes that conventional gel electrophoresis is inadequate for resolving larger DNA fragments, necessitating techniques like pulsed field gel electrophoresis or long-column liquid chromatography.

PREREQUISITES
  • Agarose gel electrophoresis principles
  • Pulsed field gel electrophoresis (PFGE) techniques
  • Understanding of DNA structure and chromosome composition
  • Knowledge of DNA migration behavior in gel matrices
NEXT STEPS
  • Research the principles and applications of pulsed field gel electrophoresis (PFGE)
  • Explore the effects of DNA entanglement on migration rates in gel electrophoresis
  • Investigate liquid chromatography techniques for separating large DNA fragments
  • Study the impact of DNA insertions on electrophoretic mobility
USEFUL FOR

Researchers, molecular biologists, and laboratory technicians involved in DNA analysis, particularly those working with large DNA fragments and chromosome studies.

nobahar
Messages
482
Reaction score
2
Hello!

If I run an agarose gel of undigested DNA of an organism with more than one chromosome, and obtain one band (slightly smeared), what does this mean? The chromosomes aren't all the same length (some might be approximately half the size of others). Is it because they all become "entangled?", they run a distance shorter (i.e. suggesting they are larger) than anyone chromosome. One could even speculate that the band has migrated a distance expected for the genome size, but this might be influenced by the above thought (since the resolution at that large a size makes it difficult to tell).

Any help appreciated,
Nobahar.
 
Biology news on Phys.org
Aha! I found pulse field gel electrophoresis, and on wiki it says the following: "DNA molecules larger than 15-20kb migrating through a gel will essentially move together in a size-independent manner."
So my undigested DNA moves at the same rate, apparently. EDIT: realized a part doesn't make sense so it is removed: I misinterpreted what it meant and in also in a way ignored what it meant.
This makes sense for why an organism with more than one chromosome shows one band. However, with a small insertion, the migration through the gell is retarded, this is not size-independent, then. If they move indpendent of size then an insertion would make no difference, so there would be no difference in migration between large DNA with or without an insertion. My results would make more sense if the DNA became 'entangled', that way insertions can be additive, which would account for a smalll inseertion making a large difference in migration, and at the same time explain (maybe) why there is only one band for an organism with multiple chromosomes.
Has anyone heard of multiple chromosomes becoming entangled?
Thanks in advance.
 
Last edited:
Basically chromosomes are just too big to be effectively separated with conventional gel electrophoresis, you need pulsed field gel electrophoresis, or possibly liquid chromatography with a really really long column length. Depending on how big the difference between chromosomes in basepairs is, you may need quite a long time to detect the differences in length (i.e. 100bp difference between chromosomes will take much longer to resolve than a difference of 20kbp)
 

Similar threads

Replies
4
Views
2K
  • · Replies 4 ·
Replies
4
Views
3K
  • · Replies 8 ·
Replies
8
Views
4K
Replies
2
Views
4K
  • · Replies 3 ·
Replies
3
Views
5K
  • · Replies 1 ·
Replies
1
Views
3K
  • · Replies 0 ·
Replies
0
Views
2K
  • · Replies 4 ·
Replies
4
Views
3K
  • · Replies 21 ·
Replies
21
Views
6K
Replies
18
Views
7K