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How to draw a DOT blot from a physical map of RFLP analysis?

Genetics as it applies to evolution, molecular biology, and medical aspects.

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How to draw a DOT blot from a physical map of RFLP analysis?

Postby supriabio » Fri Nov 27, 2009 4:08 am

hoping for some genetics help with this question

Here is the physical map of a region of DNA you are mapping by RFLP analysis:

===================
(probe)

_____|<-1.5kb->|<--2kb-->|<-1kb->|<----4Kb---->|
SITE: 1 2 3 4 5


The vertical lines with the numbers at the bottom( the numbers are beneath the vertical lines ) represent restriction sites for the enzyme EcoRI.; Sites 3 and 4 are polymorphic; the others are not. The distances between EcoRI sites are given. The extent of the probe you are using to detect the RFLP is indicated by a thick horizontal line (at top). You cut DNA from different individuals with EcoRI, separate fragments by electrophoresis, blot to a membrane, and hybridize the blot to a labeled probe. Draw a picture of what the blot will look like when the DNA comes from individuals homozygous for the following haplotypes:

HAPLOTYPE SITE 3 SITE 4
A present present
B present absent
C absent present
D absent absent

My genetics teacher is pretty terrible.I've been tearing my hair out over it since 3 days and the professor refuses to meet with me outside of class (her office hours don't coincide with hours I'm not in other classes, and even though the syllabus mentions you can make appointments to meet with her outside the assigned office hours, she's been incredibly uncooperative). We've been doing this stuff for about a week and it just hasn't sunken in yet for me.


This is what I know about how to approach this problem: All of the areas in between the restriction sites are from different individuals, right? I know that we're looking to know more about the variations in DNA sequence, and although usually knowing the object of the problem helps a lot, it doesn't seem to be helping here. I just... don't know what to do with the information I have, I guess. In class, I thought I knew what a haplotype meant, but I don't really know what it would mean to be "homozygous for the following haplotypes" followed by +/- data for letters whose origin I'm not even sure of. I guess I just need somebody to help me sort out all this information and point me in the right direction as far as how to start.

I am unable to figure out how i can represent a physical map on a DOT blot kindly help me out i really don't understand how it looks like on the paper please help me out with this i shall be great full to you. Thanks you guys.
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Postby JackBean » Fri Nov 27, 2009 6:54 am

Try to look to Wiki
http://en.wikipedia.org/wiki/Restrictio ... lymorphism

Unfortunatelly, I don't get much, what you actually need...
http://www.biolib.cz/en/main/

Cis or trans? That's what matters.
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Re: How to draw a DOT blot from a physical map of RFLP analysis?

Postby zami'87. » Fri Nov 27, 2009 10:01 am

Bands you can see are those that bound radioactive probe.The way you typed..probe can bind to region 1 and 2..
When you cut with EcoRI DNA from individual A you get two visible bands (with probe) 1.5kb and 2kb..and 2 invisible 1kb and 4kb
When you cut DNA from individual B you get again two visible 1.5kb and 2 kb and 1 invisible 1+4=5kb
When you cut DNA from individual C you get 2 visible bands 1.5kb and 2+1=3kb and 1 invisible 4kb

When you cut DNA from individual D you get 2 visible bands 1.5kb and 2+1+4=7kb and none invisible

Hope I understood your question and helped a bit
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Re: How to draw a DOT blot from a physical map of RFLP analysis?

Postby supriabio » Fri Nov 27, 2009 7:08 pm

===================
(probe)

_____|<-1.5kb->|<--2kb-->|<-1kb->|<----4Kb---->|
SITE: 1 2 3 4 5



this is the one properly aligned...


the probe is on sites 2,3,4

the distance between site 1 and 2 is 1.5kb

site 2 and 3 is 2 kb sites 3 and 4 is 1kb and sites 4 and 5 is 4 kb ....i dont see it properly aligned here

so in this case as you told me am i suppose to look at the fragments from sites 3 and 4? but i still dont understand the haplotypes part in the question please help me with this
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