By Tara L. Fulton (auth.), Beth Shapiro, Michael Hofreiter (eds.)
Research into historic DNA all started greater than 25 years in the past with the e-book of brief mitochondrial DNA series fragments from the quagga, an extinct relative of the zebra. historical DNA study quite won momentum following the discovery of PCR, which allowed thousands of copies to be made from the few last DNA molecules preserved in fossils and museum specimens. In Ancient DNA: tools and Protocols specialist researchers within the box describe the various protocols which are now prevalent to check historic DNA. those contain directions for establishing an historic DNA laboratory, extraction protocols for a variety of diverse substrates, information of laboratory concepts together with PCR and NGS library training, and recommendations for applicable analytical ways to make feel of the sequences bought. Written within the hugely profitable Methods in Molecular Biology™ sequence layout, chapters comprise introductions to their respective subject matters, lists of the mandatory fabrics and reagents, step by step, simply reproducible laboratory protocols, and key pointers on troubleshooting and averting recognized pitfalls.
Authoritative and sensible, Ancient DNA: tools and Protocols seeks to assist scientists within the extra examine of historical DNA and the methodological techniques in historic research.
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Extra resources for Ancient DNA: Methods and Protocols
10. 11. Cooper A (2002) Flight of the dodo. Science 295:1683 Johnson KP, Clayton DH, Dumbacher JP, Fleischer RC (2010) The flight of the passenger pigeon: phylogenetics and biogeographic history of an extinct species. Mol Phylogenet Evol 57:455–458 Stamatakis A (2006) RAxML-VI-HPC: maximum likelihood-based phylogenetic analyses with thousands of taxa and mixed models. Bioinformatics 22:2688–2690 Huelsenbeck JP, Ronquist F (2001) MRBAYES: Bayesian inference of phylogenetic trees. Bioinformatics 17:754–755 Ronquist F, Huelsenbeck JP (2003) MrBayes3: Bayesian phylogenetic inference under mixed models.
Mix well (see Note 3). 2. Add a suitable amount of digestion buffer to the sample (see Note 3). Vortex briefly to ensure that any pelleted powder is homogenised in the solution. Incubate the sample plus buffer overnight at 55°C with gentle agitation. 3. Keratinous samples may not fully digest after this incubation. If full digestion is required, add an additional 40 mL 1 M DTT solution and 100 mL proteinase K solution to the mixture, vortex briefly, and return to incubation with agitation for at least 1 more hour.
It is wise to resuspend the silica pellet by placing the tip of the pipette at the edge of the pellet in the base of the tube and pipetting up and down slowly. Be careful not to allow the solution to bleed over the edge of the tube. While 50 μL of silica is sufficient for more than 10 μg of DNA, the silica clearly gets “clogged” with other polar molecules. Using less than 50 μL is therefore not advised. However, in our experience, increasing the amount of silica above 50 μL has not been shown to yield quantitatively more DNA.