If you extracted DNA from liver cells, rather than muscle tissue,
would you expect a similar result? explain your answer.
Our group disagrees on whether we would more or less. I feel that we
would get the same amount. The only difference I can see is the
amount of mRNA that the cells would contain. Am I correct in this
assumption.We also disagree on what does "denature the probe" actually
mean. Can you help settle these arguments
Thanx |
Request for Question Clarification by
synarchy-ga
on
25 Apr 2003 06:16 PDT
Hello - what type of experiment are you doing on the DNA that was
extracted from muscle? While xarqi-ga is quite right in saying that
the content of nuclear DNA will be same (on a per cell basis), with
possibly some variation in the content of mitochondrial DNA, there can
be other differences in the composition and conformation of the DNA
that are tissue specific - without more information regarding the
particular experiment, it is difficult to know.
synarchy
|
Clarification of Question by
marmite-ga
on
26 Apr 2003 15:09 PDT
WE extracted DNA from muscle tissue. One of the questions that was
asked in relation to the experment was if we used liver tissue would
we get a similar result. The experiment was extracting
DNA,restriction digestion using Eco Rl adding a probe and doing a
southern blot from the gel and visualising it with a dig kit probe.
The experiment was not sucessful as our blot did not produce any
colour but the initial gel did show a band where we should have one. I
hope this helps
Thanx
|
Request for Question Clarification by
synarchy-ga
on
27 Apr 2003 18:40 PDT
What was the probe to?
|
Clarification of Question by
marmite-ga
on
28 Apr 2003 15:11 PDT
A DIG-labelled DNA probe (supplied to the students) was hybridised to
the membrane which was prepared by capilliary blotting (southern
Blot)The labelled probe to complementry sequences on the membrane and
then detected using a DIG colour reaction. I trust this is what you
mean by what was the probe to?
Thanx
|
Request for Question Clarification by
synarchy-ga
on
28 Apr 2003 17:10 PDT
What were the "complementary sequences"? ie - to what gene/repeat
element/etc was the probe directed against?
The only reason that I ask is that it is possible for
probes/restriction enzymes to not bind to DNA sequences that have been
modified by methylation - if the target of the probe was known to be
differentially expressed in muscle tissue versus liver tissue, then
the blot may not produce the same results if the DNA is modified, and
if that interferes with either the probing or cutting processes.
From your descriptions so far, I would guess that this is an
unnecessarily complex issue for what is likely an introductory
molecular biology class. The quantity of DNA should be the same.
|
Clarification of Question by
marmite-ga
on
29 Apr 2003 14:04 PDT
I reply I don not know. We extracted the DNA, made the probe, cut the
DNA and run a gel then transferred the fragments from the gel to the
nylon membrane anf then hybridized with the probe. All we where
looking for was a colour change on the membrane.
|
