To complement the PrediCtoR software being developed in JRA1, JRA2 will develop a new protocol that will enable users to estimate the ancient DNA preservation status of rare and unique bone specimens far more accurately than is possible with current protocols. PCR of extracted DNA is by far the best current method to test for the survival of ancient DNA, but its main drawback is that is always destructive. JRA2 aims to devise a new protocol/tool that correlates with PCR’s levels of success with only minimal and hardly visible destruction of specimens. By comparing PCR success of selected bone samples with data gathered from various state-of-the-art screening techniques JRA2 will identify a low-cost technique that best predicts the presence of usable DNA.
- Statistical evaluation of DNA preservation/degradation factors
- Developing non-destructive or minimally-destructive techniques (tests to include: UV- or IR-spectroscopy, immunohistochemistry, immunochemistry of decalcified bone, protein gel electrophoresis, surface antibody staining, laser microscopy).
- Based on the known PCR success from specimens (resulting from JRA3), define an optimal non-destructive technique with results that correlate with increased preservation of DNA (without destroying precious museum specimens). Such a refined tool for predicting good DNA recovery will greatly enhance PCR success and lead to the relaxing of protection policies and liberate more fossil/bone specimens for DNA studies
- If the optimal tool developed is portable, it will be rolled-out to Users and collections managers to evaluate correctly the DNA preservation of new finds on location
The new system will enable Users to estimate the DNA preservation status of rare collections specimens more accurately. PCR of extracted DNA is the best current method to test for the survival of ancient DNA; its main drawback is that is always destructive. JRA2 aims to devise a new method that correlates with the PCR levels of success without the destruction of specimens. By comparing PCR success of selected bone samples with data gathered from various state-of-the-art screening techniques JRA2 will identify the technique that best predicts DNA preservation. This will become an essential tool for curators and bone collection Users as it will both facilitate more efficient specimen selection and the avoidance of specimens with poor preservation profiles.
JRA2 will combine the taphonomic knowledge obtained for all techniques in order to find a simple and easily applicable screening technique that can potentially be adopted by Beneficiaries and beyond without the need for highly specialist laboratories. The final aim of JRA2 is to develop a DNA screening device that will allow the collection manager to predict the utility of freshly excavated bones to yield usable DNA on–site with a non-destructive tool.
Evaluating DNA preservation/degradation factors
Generate and deliver statistically-validated analysis that reveals the key and easily measurable factors that predict good preservation of DNA in archived bone specimens. Based on this, JRA2 will include these factors in PrediCtoR. This will improve the Web-based retrieval of the individual history of a find and thus the ancient DNA survival prediction. The results will also yield a theoretical basis for developing the most appropriate pre-dissemination screening technique.
Identifying the optimal screening technique and development of a screening device/protocol
Empirically determine a screening technique which highly correlates with successful ancient DNA preservation in bones. Depending on predictive results JRA2 will develop a low-cost, pilot screening device. There is a risk of that the favourable correlations will only result with high cost technology (isotope analysis). JRA2 will address this by focusing the tests on easy to apply techniques, such as UV- or IR spectroscopy that, in combination with PrediCtoR, could potentially provide the results that correlate with DNA yield.
Testing the screening device/protocol
In 2013, the established procedures/protocols will be tested. Therefore, with a limited number of bone samples will be acquired from museums. Within the framework of a blind test, DNA results will be compared to the predictions made by museum curators usingPrediCtoR and the selected screening method/device.
JRA2 investigated creation of DNA libraries to reduce the need to re-sample rare museum specimens by effectively immortalizing one DNA sample. Ancient skeletal material was used as the model as the DNA is often both highly degraded and in demand from users.
JRA2 has developed, tested, and optimized new protocols which used in combination with next generation sequencing technologies can be applicable to the degraded DNA found in museum and herbarium samples. In particular, protocols for the creation of DNA libraries out of both skeletal museum specimens and soil embedded archaeological remains and in-solution capture enrichment (capturing genomic regions of interest from a DNA sample prior to sequencing using magnetic beads) were investigated.
- Development of a library preparation protocol adopted to highly degraded DNA (Deliverable 5.4 of the SYNTHESYS2 project)
- Protocol delivered adopted to the characteristics of hominin aDNA (Deliverable 5.5)
- Protocol for the immortalization of aDNA libraries (Deliverable 5.6)
- Development of a showcase capture-NGS protocol (Deliverable 5.7)
- Report on the success rate of mtCapture protocols applied to various different museum and archaeological specimens (Deliverable 5.8)
- Final protocol for mitochondrial and nuclear capture enrichment (Deliverable 5.9)
There are no planned events at this stage.
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