Dec 11, 2024
Version 1
HWT buffer V.1
- 1Max Planck Institute for Evolutionary Anthropology
- MPI EVA Ancient DNA Core Unit
Document Citation: Anna Schmidt, Anya Patova, Matthias Meyer 2024. HWT buffer. protocols.io https://dx.doi.org/10.17504/protocols.io.x54v92kkml3e/v1
License: This is an open access document distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Created: July 24, 2024
Last Modified: December 11, 2024
Document Integer ID: 103970
Funders Acknowledgements:
Max Planck Society
Grant ID: -
Abstract
Protocol for the preparation of HWT buffer (Maricic et al. 2010, Fu et al. 2013), which is used in hybridization capture by the Ancient DNA Core Unit of the MPI-EVA.
References
Fu, Q., Meyer, M., Gao, X., Stenzel, U., Burbano, H.A., Kelso, J., Pääbo, S. (2013) DNA analysis of an early modern human from Tianyuan Cave, China. Proc Natl Acad Sci U S A. 110(6):2223-7. doi: 10.1073/pnas.1221359110
Maricic, T., Whitten, M., Pääbo, S. (2010) Multiplexed DNA sequence capture of mitochondrial genomes using PCR products. PLoS One, 16;5(11):e14004. doi: 10.1371/journal.pone.0014004
Note
This protocol describes the preparation of 500 ml buffer.
Materials
| Reagent/consumable | Supplier | Catalogue number | |
| Reagents | |||
| Water | Sigma Aldrich/Merck | 1153332500 | |
| 10x AmpliTaq Gold buffer without Mg | Life Technologies | 4379874 | |
| Tween-20 | Thermo Fisher Scientific | 11417160 | |
| Consumables | |||
| Square media bottle 500 ml | VWR | 391-0630 | |
| 50 ml serological pipette | Corning BV | 357550 | |
Equipment
- Serological pipette controller (e.g. battery-powered pipetting aid ROTILABO, cat. no. TC16.1)
Protocol
1. Prepare the buffer in a 500 ml square media bottle by adding the following reagents. Use the glass pipette for transfer of large volumes (> 1 ml). Mix reagents by shaking the bottle.
| Reagent | Volume | Final concentration in reaction | |
| Water | 449.9 mL | ||
| 10x AmpliTaq Gold buffer (w/o Mg) | 50 mL | 1x | |
| Tween-20 | 0.1 mL | 0.02% |
Note
[Labeling]
Label the bottle with the buffer name, batch ID, date and the initials of the person who prepared the buffer. Include lot number of AmpliTaq Gold buffer.
Attention: Every single bottle prepared at the same day gets a new batch ID. Name the batches with Roman numerals (e.g. batch I, batch II, etc.)
2. Store the buffer at room temperature until used. Shelf life is at least one year from preparation.
Note
[Documentation]
Note the lot numbers, date and initials written on the reagents used for buffer preparation in Labfolder (orange fields).