Try Alan Cooper, Morten Allentoft or Thomas Gilbert.

Thanks Federico.

Qiaomei Fu- Kindly refer this

1. Oskam CL, Haile J, McLay E, Rigby P, Allentoft ME, Olsen M, Gilbert MT, Holdaway RN, Willerslev E, Bunce M, (2010) Fossil avian eggshell preserves ancient DNA. Proc. R. Soc. B. 277,1991–2000.

Try Brian Kemp and also I would be interested. I normally work with human skeletal material but it would be fun to try! I am in the Genomics Core at OHSU in Portland, OR.

Try me! My lab is specialized in aDNA work from geological samples and we have also extracted aDNA from similar kind of Ca carbonate-rich samples such as marine carbonate oozes, stalagmites and corals. The problem with this kind of material is that released DNA during extraction will reabsorb completely  to the mineral matrix within seconds so you won't have a chance to keep it in solution using standard methods. We have an unpublished method that minimizes this reabsorption so it dramatically increases the field,

Both of us.......one on each side of the country to validate.......

Typo: I meant yield not field

Interesting, Marco - my experience and expertise is human skeletal and dental remains as old as 7000 BP and microbial sequencing recovered from difficult environmental samples.  Hmm, and RNA from bone so I would say calcified tissues.....It would be a great opportunity to collaborate..........

But it seems that you are very successful in extracting the DNA from those samples. Correct? I am actually moving to Curtin University, Pert Australia next month. My lab will probably not ready before March because shipping my lab from here takes 3 months.

Cooper and Gilbert are great choices if you can get one of them.

Yes - I have been successful with those samples - but you have experience with the issues of ca carbonate rich so I would suggest collaboration! Of course, its all up to the PI, Sonal, on who, what, and where! But - I would like to stay in contact with you for future collaborations? Congrats on the new lab!

Charlotte Oskam has plenty of experience dealing with DNA in egg shell so I'd certainly try her.  Mike Bunce at Curtin will know how to find her, if she is not still working with him.

@ Amiee and Marco- We have done confocal microscopy on these eggshells and observed DNA hotspots as suggested by Oskam et al 2010. We have tried extracting DNA but were unsuccessful. We need some expertise  to extract aDNA from these eggshells as they are gateway to a lot of information. As they are from Upper Paleolithic sites from India, so probably DNA is degraded. 

There are two issues - first, extraction that will maximize recovery of potentially non-intact fragmented and low copy number DNA but minimize co-extracted inhibitors and then PCR and sequencing of small amplicons to accommodate degraded DNA. You were unsuccessful using their protocol? Did qPCR indicate no DNA? Are you sure about target sequence? How did you quant your extracted DNA?

I used to make all my own reagents and buffers and then use column or P:C:I organic cleanup and precipitation. However, I have to say that now I use the Qiagen DNA Investigator kit but modify the length of digestion to a week. I had great success with this kit on my most recent aDNA project.

We have tried the procedure as described in Oskam CL, Haile J, McLay E, Rigby P, Allentoft ME, Olsen M, Gilbert MT, Holdaway RN, Willerslev E, Bunce M, (2010) Fossil avian eggshell preserves ancient DNA. Proc. R. Soc. B. 277,1991–2000.

We quantified DNA using bioanalyzer.

That sounds like a neat project! If I had a lab I would be all over that.  Well, I do have a lab, sort of. Message me and we can talk in more detail.

Hi Sonal,

I would try a nanodrop rather than bioanalysis.....The Agilent isn't going to be as accurate and sensitive for the low concentration samples you have. The nano drop won't be if either if you are down around 10ng/ul (which you probably are). I do have access to a full genomics [email protected]. 

Also - with LCN samples nuclear DNA can be undetectable but there is enough mtDNA. 

Please let me know of anyone doing/ willing to do similar work on plant fossils in india.

@ Avik- Birbal Sahini institute of Paleobotany.

@ Sonal: they primarily do paleobotanical description, not palogenetics