jamestoon1

Supposed i have 2 polymorphisms, SNPA and SNPB.
Individuals who carry the wildtype genotype for SNPA also carry the wildtype genotype for SNPB.
Individuals who carry the heterozygous genotype for SNPA also carry the heterozygous genotype for SNPB.

Individuals who carry the mutant genotype for SNPA also carry the mutant genotype for SNPB.

It is pretty obvious that the two polymorphisms are in complete linkage disequilibrium, right?
So the D' should equal to 1.

But, how do I calculate that?

I found a web-based software which seems to be easy to use, but I don't know what number should I key in onto each cell --> http://www.oege.org/software/cubex/

Now supposed I have 1000 wildtype, 800 heterozygous and 600 mutants for both polymorphisms, what number shall I put in the cells?

I tried the following (based on the frequencies of my genotypes):
2000---1800---1600
1800---1600---1400
1600---1400---1200

But I got a  D' value of -0.011 instead of 1.

Can anyone help?

You may recommend other softwares or simple methods for calculating the D' and r2.

Thanks in advance.

Today I isolated RNA from 16 samples and quantified the RNA using spectrophotometers.
15 of the samples had A260/280 ratio between 1.9 and 3.5. I think the range was still acceptable although 3.5 is actually quite high.
But...the remaining one sample had an A260/280 ratio of about 30!
I checked with 2 spectrophotometers and both gave me the same reading.
What does this absurdly high A260/280 ratio mean?

My DNA and RNA isolation kits require me to dilute some of the buffers with absolute ethanol.
I added denatured ethanol to all the necessary buffers, then only I came to realize that we should use non-denatured ethanol for molecular biology applications because denatured ethanol contains impurities.
What should I do now?
Can I proceed with DNA and RNA isolation using the reagents?
If not, my supervisor is going to kill me OMG!!!

Hi.

I isolated RNA from human tissue samples using RNeasy.
I electrophoresed the RNA obtained and this is what I got:
 is my rna degraded.JPG   16.41K   79 downloads
P.S. 100bp DNA ladder was used instead of RNA ladder because we don't have RNA ladder in our lab.

From the gel image, two clear bands can be seen (supposed to be 18S and 28S rRNA).
But there are also some smaller bands down there.
Does this indicate RNA degradation or something else?

P.S. The tissue samples were preserved in RNAlater for 1 month before RNA isolation.

P.S. We don't have Bioanalyzer in the lab, so I cannot analyse it in the instrument.

P.S. I did not check the purity and concentration of the RNA because the only spectrophotometer in our lab has been send for repair.

When isolating DNA from cells in suspension, most if not all the protocols I have come across recommend centrifugating the suspension at the very beginning to obtain a cell pellet, followed by resuspension of the pellet in either water or PBS or other buffers.

May I know what is the purpose of obtaining this cell pellet?

Can we just vortex the suspension and use it directly without pelleting the cells?