Jamey Tolman and Dr. Scott Woodward, Microbiology
Recent advances in DNA sequencing technology have allowed us to gain knowledge about the ancient populations of the world. A team of BYU students and faculty has excavated a group of bodies, dating from 200 B.C. to 400 A.D., found in the Fag El Gamous cemetery in Egypt. A major observation from the excavations has been that the earlier, or lower burials, in the cemetery are buried with their heads to the east. As you move up in the cemetery, to more recent dates, a distinct change occurs in the orientation of the burials. The bodies are placed with their heads to the west. We believe that this change in burial patterns in this cemetery were directly affected by changes in the population of people that resided in the same area. It is proposed that these changes can be identified by the genetic signatures present in the DNA of the ancient population. Previous genetic analyses of samples taken from these people have revealed clusters of family based burials. Through further genetic analysis of the mitochondrial DNA (mtDNA) of these people, we would like to be able to answer a significant anthropological question concerning the origin of these changes in burial customs.
There are many different possibilities as to what could have caused this change in burial patterns. One possibility is that the population that first existed died out for some reason around the date indicated, and a completely new population moved in along with their new burial customs. This would be shown by drastic changes in comparisons of DNA sequences of those sampled. DNA types reflecting the older population would not be found among the most recent samples. Another possibility is that a new population migrated into the area, brought their different burial customs, mixed with the already present population, and together they resided in the area. This would be shown by a significant change in the patterns of DNA, but there would still be a distinct continuation of sequences from the first population. A third possibility is that there was no second population. Some type of change in philosophical and/or religious practices caused these people to change their burial customs. This would be evident by no significant change in DNA patterns throughout the population over the time studied.
In our study of this ancient population, the displacement loop (D-loop) of the mitochondrial DNA (mtDNA) was used. It is a circular sequence of DNA located within each mitochondria of the cell. Since each cell contains multiple copies of the mitochondria, there is a higher chance that those sequences of DNA have survived degradation. This is important in amplification of ancient DNA, because its isolation is naturally more difficult than contemporary DNA. Because it is a non-coding region, mtDNA sustains a higher rate of polymorphisms and is usually preferred over nucleic DNA.
Generally tooth and bone extractions have proved to be the most effective in isolating ancient DNA. By drilling into the tooth or bone for the DNA, these types of extractions aid in preventing contamination of the sample. For this project, tooth extractions in a lysis buffer were used. Negative controls with every extraction and later reactions were used to detect the presence of any contamination or false positives.
In this project, a series of polymerase chain reactions (PCR) with specific primer sets were used for amplification of the DNA. Since ancient DNA becomes very degraded over time, the first is an attempt to repair nicks and breaks in the DNA that has been extracted. A normal PCR is then used to amplify the sequence of DNA that is to be studied. Through automated sequencing, the specific sequence of DNA was determined, and can be compared with associated sequences by relationship matrices and/or absolute distance matrices.
The DNA isolation procedures were performed on twelve different samples. Working sequences from three distinct samples were obtained. Two samples, SW43 and SW46, yielded sequences of approximately 200 base pairs apiece in hyper variable region 2 of the mtDNA, and the resulting over-lapping sequence consisted of 100 base pairs. These samples were congruent to each other throughout the 100 base pairs, but they differed from Anderson, a standard sequence, by two polymorphisms. When compared to corresponding modern sequences from other populations around the world, there were many that were similar. A larger base pair sequence will be needed to reach a more concrete definition of exactly where these sequences have ended up in the modern population. Another sample, SW38, gave some very unexpected results. A 500 base pair sequence was obtained with the same primer set used with SW43 and SW46. Comparisons of this segment of DNA to the sequences of other organisms throughout the world showed that segments of this DNA sequence most closely matched that of Halobacterium, a saltloving specie of Archaea. Further amplification and analysis will need to be done to confirm these results.
Although there is still much more work to be done in this project, I have really been pleased with how the research has gone so far. I have learned much about the scientific method, correct principles of research and dedication to a project that takes consistent hard work and repeated trial and error. Findings and results from this research will be of great benefit to future projects.