Genotype of snp at terc and other loci.

Good advice, thanks!

Does anyone have any consideration regarding environmental factors?

In fact, there are several factors (as you said is overwhelming) -- that's right!

But, in case of telomere shortening and having deterioriating effect on telomere shortening are the following factors that we can take into account for the time being, which is still controversial as suggested by the papers I am following: like BMI (Basal Metabolic Index or Body Mass Index), habit of smoking, lipid profie of an individual (for example: higher level of LDL and lower level of HDL in general), age, sex (since female tends to have longer telomere length compared to male counterparts), habit of physical exericise (whether or not that subjects are practicing physical exercise). You can adjust with these parameters!

Oh sorry, in fact, physical exercise have positive effect on maitaining the optimal or good length of telomere. ;-) Sorry for that in above comment.

Other environmental factors includes: subject to the sun light or UV exposure, work pressure, diet and foods. But, I think these factors are not supposed to be adjusted while doing statistical calculations in this case.

Also include anti-inflammatory medications that play a role in telomere dynamics, as well as previous/chronic diseases (especially cancer)

You might also want to look at paternal and maternal age, if you have that information.

Thank you all for the good advice.

I have a question to Sarah: do you have a paper that evaluated the correlation between telomere length and the age of the parents? If you do, could you send it to me? I am asking because I have not heard from this potential correlation before...

Can I check with you on your qPCR method, which protocol are you using? I am doing similar work but having difficulty getting it to work.

Sarah is right. Yeah, if you have maternal age, it would give more clear picture. Because one study based on the twins sister with premenopause and postmenopause have found that there is a strong relationship between the level of estrogen and the telomere length. Potsmenopausal women doesn't have significant difference in the length of telomere than with the control or something. I am not quite sure of this. But, check this one too.

Ene choo Tan: Can you make it clear whether you are measuring the relative telomere length or the absolute telomere length. Usually, the primer used in the real-time PCR for the telomere plays a crucial role. Usually, most of the studies conducted till now, I have found they all are using the primer desiged by Cawthon. Another thing -- the thermal profile, concentration of the primer and the DNA template you are using might be causing problem. Follow cawthon paper. It will give you some insight. All the best. :-)

We tried the Cawthon conditions and also others, and both relative and absolute ratios. The data doesn't seem to correlate with TRF measurement. Would like to seek your opinion if you can email me at [email protected] Thanks!

Fernando, here is a recent publication regarding paternal age and telomere length:

Proc Natl Acad Sci U S A. 2012 Jun 11. [Epub ahead of print]

Delayed paternal age of reproduction in humans is associated with longer telomeres across two generations of descendants.

Eisenberg DT, Hayes MG, Kuzawa CW.

Thanks for the paper Sarah!

Ene: we are following Cawthon's protocol with some modifications. We are not using the additives they proposed (DTT and DMSO) since, in our experiments, they did not changed the amplification efficiency. Moreover, our reactions are of 10 ul (instead of 20), but we did not change the DNA nor the primers concentrations. We work with sybr green (Applied Biosystems) in a 1:2 dilution rate (so we use 5 ul for each reaction). The cycling conditions are the same as Cawthon's.

Using this protocol we got very satisfatory primer efficiency rates. If you want more details, please let me know.

In case you would like to e-mail me: [email protected]

Thank you guys for your help!

Here's a discussion of the influence of maternal age in the case of Down's syndrome:

Telomere length is associated with types of chromosome 21 nondisjunction: a new insight into the maternal age effect on Down syndrome birth.

Ghosh S, Feingold E, Chakraborty S, Dey SK.

Hum Genet. 2010 Apr;127(4):403-9.

PMID: 20063167

do not forget the potential confounding factors from the genetic endpoints. I would select one ethnicity. From my experience you also want to harmonize the experiment to avoid batch effect on the telomere length side.