A Comparative Study of Sequences Derived From Mitochondrial DNA Databases Applied To the Question Of the Origin Of Modern Humans

Transcription

1 Literature Review A Comparative Study of Sequences Derived From Mitochondrial DNA Databases Applied To the Question Of the Origin Of Modern Humans Kara B. Babrowski Abstract Molecular data is increasingly used in the study of the origin of modern humans. Several molecular markers are available for study, however the most widely used is mitochondrial DNA (mtdna). In 1987 Rebecca Cann, Allan Wilson and Mark Stoneking published the seminal paper demonstrating the power of mtdna as a tool for analyzing human origins. Since then, a large body of literature has been published disputing their findings. Most commonly cited points of disagreement concern sample size, sample sources, molecular markers and lack of adequate statistical tests. In this study, human mtdna diversity is considered. A large sample of mtdna sequences from the D-loop region of individuals from diverse origins will be selected from MitBase (a database available on the World Wide Web). It is the aim of this research to use this data to reanalyze Cann, Wilson and Stoneking's work. Through the use of a separate data set, the attempt will be made to generate similar conclusions as previous studies such as Cann, Wilson and Stoneking's. Quick Index [Mitochondrial DNA Databases] [LIT. REVIEW] [DATA] [ANALYSIS] [CONCLUSION] [REF. CITED] Proceed [11/30/2007 1:19:57 PM]

2 Mitochondrial DNA Databases A Comparative Study of Sequences Derived From Mitochondrial DNA Databases Applied To the Question Of the Origin Of Modern Humans Kara B. Babrowski It has been nearly twelve years since the scientific community saw the publication of the first valid research regarding mitochondrial DNA in conjunction with human evolution. Since this time, there has been an exponential growth in publications on this topic, due in particular to the innovation of the Polymerase Chain Reaction by Kary Mullis in the 1980's. This revolutionary technique provided a method by which tiny portions of DNA can be amplified, allowing the sequences of fossils as well as modern humans to be determined. The continuing analysis of the mitochondrial genome of antiquated skeletons along with contemporary humans is certain to provide a view into our evolutionary past. Although many scientists are attempting to link these old fossils to modern humans, there are many different methods to meet such an end. Some of the research being conducted in this discipline is mathematical as it is based upon previous sequences, while some deals with the sequencing of different mitochondrial genomes. There is a tremendous body of information available for many sequences, yet there have not been any conclusive findings. This has resulted in little effort being put forth to look at this research from a macroscopic point of view. Not all paleoanthropologists have fully accepted the validity of the sort of research presented in this paper which is another reason why the synthesis of the current data is not complete. Through the digestion and careful analysis to follow, it is the goal of this paper to carefully analyze the current research and lend credence to the conclusions presented in the various journal articles. This paper will also look at one current application of mitochondrial DNA research as it relates to the Out of Africa Model of the origins of modern Homo sapiens. This analysis relies on the use of the MitBase, an Internet database containing relevant mitochondrial DNA sequences. The sequences gleaned from MitBase were downloaded from the Internet and then copied into MacVector, which serves as an alignment program. These sequences were then aligned and each sequence individually compared to the entire body of entered sequences. Comparisons of the results achieved through this technique will be made to the current literature in order to evaluate the extent of their agreement to the most recent hypothesis of the origins of modern human. With the availability of databases on the Internet to the public, it has been possible to perform original tests on previously published sequences. A result of this analysis is to demonstrate the practical application of Internet sequence databases. This will be illustrated thorough the implementation of the computer program MacVector to show there is more difference within African D-loop mitochondrial DNA sequences than exists within (1 of 3) [11/30/2007 1:19:59 PM]

3 Mitochondrial DNA Databases other population including Indonesia and Native American Indians. Upon investigating all of the material at hand, and applying a multidisciplinary perspective to mitochondrial DNA, it is certain to be concluded that the direction of mitochondrial DNA research must be pushed in this direction. The above-mentioned molecular approaches have enjoyed limited application which, when used, prove to be more conclusive than the traditional skeletal analyses. These techniques must be used to help put the Multiregional Hypothesis to rest in favor of the Out of Africa Hypothesis. The new frontier of "molecular anthropology" began in the 1970's with the work of Vincent Sarich, Masatoshi Nei and Arun Roychoudhury (1974). It was at this time that Nei and Roychoudhury published molecular data showing Europeans and Asians are together both more closely related to one another than either is to the African population. Nei and Roychoudhury concluded the European-Asian population had a distinctly different genetic make-up than the African population. From this, they construed the Africans must be an older genetic group. At that time, they calculated a split of 130,000 years between Africans and the European-Asian populations. Though this was the first evidence presented of Africans being molecularly separate and a great deal older, technology was not yet available to test the genetic make-up of the two populations. Approximately thirteen years later, technology had advanced sufficiently to cause this hotly debated subject to again come to the fore with the proposition by Cann, Wilson and Stoneking (1987) that modern humans originated out of Africa approximately 200,000 years ago. This work was based on the molecular analysis of mitochondrial DNA sequences. The first documented use of mitochondrial DNA was cited in 1983 with the work of Douglas Wallace. Data from mtdna analyses led them to conclude an Asian origin of modern humans was as likely as an African origin of modern humans. However, after additional research they later agreed with Cann, Wilson and Stoneking. and concluded the origins of modern humans to be in Africa. Mitochondrial DNA is the most commonly used molecular tool being used in the attempt to understand our ancient origins. This popularity can be attributed in part to the fact that mtdna is maternally inherited; thus the genetic trees relating mtdna sequences are readily interpreted as genealogies reflecting the maternal history of our species. In addition, mtdna is commonly used because it appears to evolve rapidly. This means many differences arise even among mtdna's from closely related populations. The D-Loop region is the most commonly used region in mitochondrial DNA studies based upon the idea that it is a highly conserved region among the species. This aspect makes it ideal for study because it is a region of the DNA that is least likely to endure mutations and thus remain constant throughout populations. Though there has been a great deal of evidence relating the origin of modern humans (2 of 3) [11/30/2007 1:19:59 PM]

4 Mitochondrial DNA Databases back to Africa, several opponents of this theory exist. Milford Wolpoff contends that the distinctive regional populations of Homo erectus found throughout Africa, Asia and Europe each evolved independently into modern humans at approximately the same time. There are several supporters of this theory, but a lack of molecular evidence is widely regarded to weaken this argument. In classical evolutionary theory, it is widely held that independent evolution does not occur. This means pockets of geographically distinct populations do not hold the capacity to generate the same changes necessary to be considered the same species after a given period of isolation. Based upon the chance involved in evolution, independent evolution is entirely unlikely between two groups, let along populations spanning three continents. Through the analysis of mtdna it is likely that in the near future our ancient history will be unraveled, and more data is certain to be presented to further intensify the Out of Africa Model of the origin of modern humans. Proceed (3 of 3) [11/30/2007 1:19:59 PM]

5 LITERATURE REVIEW LITERATURE REVIEW Prior to 1987 the geographical location of the origin of modern humans (Homo sapiens sapiens) was subject to a great deal of controversy, dividing not only the anthropological community, but the general public as well. A research team headed by Rebecca Cann investigated this mystery and culminated their research by publishing a paper in Nature (Cann et al.:1987: 325). This paper is regarded as unique because it presented the first empirical evidence for the origin of humans in Africa, formulating and testing hypothesis related to anthropology in an objective manner. Commonly referred to as the "Out of Africa Model" or the "Eve Hypothesis", the ideas presented in this research are today the most widely held views about modern human origins among physical anthropologists. There still remain a few adherents to the hypothesis that modern humans evolved in several pockets throughout Asia and Europe. The Cann et al. study initiated this current hypothesis though it is actually composed from older theories on African origins. The data presented by Cann et al. (1987) has withstood the test of time, and is still very convincing even as it is riddled with several oversights. Since the 1987 publication date, there has been a great deal of technologic advancement in instrumentation as well as conceptual insights that can be used for an analysis of this classification. As previously discussed, critics of this study encountered several problems while analyzing the data. Since the initial criticism, many other aspects of this study can be questioned regarding the validity of this research given current methods and instrumentation that were not available twelve years ago. This study analyzed mitochondrial DNA from a sample size of 145 individuals. An average of 370 restriction sites were mapped per mtdna molecule, and these maps were compared to each other. A phylogenetic tree was then created from those maps and it indicated an African origin of humans, as the African populations were more deeply rooted in the generated phylogenetic tree than the other sampled populations (Cann et al.:325:1987). Though the rough sketch of this paper is very convincing, and has yet to be disproved, the fine-tuned aspects of this paper demand attention. Today, it is much easier to be critical of published research related to the genetic code of nearly all organisms. This is because all sequences ever carried out using public funding are available to the public via the World Wide Web. Those published sequences are commonly referred to as reference sequences and make it much easier to compare recent work to previously sequenced mtdna. Most researchers engaged in molecular evolutionary studies today refer to GenBank in part or whole to obtain information on their organism of study. The exponential growth of GenBank has made it possible for research to be completed at a rapid rate given the availability of information. GenBank makes available whole genome sequences as well as segments of the genome. Though many organisms have a niche in this database, the Homo sapiens sequences maintain the number one spot in number of entries. Much of the database of human sequences is comprised of mtdna sequences, which is of (1 of 5) [11/30/2007 1:20:01 PM]

6 LITERATURE REVIEW particular relevance to human evolution and is the molecule used in the Cann et al. study (1987). GenBank is a conglomerate that is managed by the National Library of Medicine through the National Institutes of Health. It is a thoroughly organized database that has several query systems from which one can access the data. A Medline accession number follows each entry, and most of the sequences also refer to the original article in which the data appeared. This is very useful when one wishes to compare previous research with the research they are performing on the same sequence. Though this database it often slow in retrieving information, as is a common difficulty with many databases on the Internet, the body of information it contains is essential to further not only the studies in human evolution, but science as a whole (Benson et al.1998:26). It is frustrating, however, when one notes to this date, it does not appear as though any relevant anthropological publications have been made as a result of this database that has been accessible since This again serves to underscore how important it is that people take advantage of these resources. John Avise has provided a vast body of information necessary for the analysis and interpretation of mitochondrial DNA. Avise's book is often cited in evolutionary studies and is viewed as a tool for use in practical applications to mitochondrial DNA. Information regarding analysis of the generated data is extrapolated from this book. It is a very expansive volume, encompassing every aspect of molecular markers in the study of evolution. Of particular use to this study is the introduction of molecular markers, which helps generate a working knowledge of the implementation of such information. The greatest use of this book is for the information that is supplied regarding how to implement molecular markers in the lab, and subsequently how to analyze the data obtained. Information is also provided regarding the calibration of the molecular clock, and procedures for phylogeny reconstruction (Avise: 1994). Though Avise is thorough in his treatment of the subject, clearly it would be impossible to create an encyclopedic work in which all aspects were fully discussed. The heavy criticism brought about by the Cann et al paper was addressed by Linda Vigilant and co-workers (1991), including Allan Wilson and Mark Stoneking who were authors of the original 1987 paper. The goal of this critique was to address several of the problems in the Cann et al. study by using a different portion of the mtdna (hypervariable regions I and II), along with different methodology. This alternative methodology was more concise and in accordance with the technological advancements that had occurred in the four and a half years since the 1987 study. Phylogenetic analysis is an important aspect of any evolutionary study, and as in the case of most analyses, there are several theories as well as methods behind the construction. Cann et al. (1987) employed the midpoint method in the analysis; a method in which the construction did not account for the apparent diversity of rates of mtdna mutation within each population. Accounting for this shortfall, Vigilant et al. implemented outgroup analysis using maximum parsimony via the PAUP (2 of 5) [11/30/2007 1:20:01 PM]

7 LITERATURE REVIEW program (Swofford 1991) in which the obvious outgroup in human evolutionary studies would be a chimpanzee. Presently, outgroup analysis is the only method employed in phylogenetics. The Cann et al. study was not able to implement this given the lack of a published chimpanzee sequence in the literature at the time of publication. Since this study, Nei and Takezaki (1996) have taken a different approach towards the phylogenetic analysis of human origins. Using appropriate statistical precautions, the chimpanzee as an outgroup and a different molecular marker (microsatellites from nuclear DNA), Nei and Takezaki have potentially generated a method that can be used in future phylogenetic analysis, regardless of the marker used to generate the actual data for analysis. Given a different body of information as well as different method of analysis, the conclusion still supported the hypothesis that modern Homo sapiens originated out of Africa (Vigilant et al.: 1991:253). Considering the importance of the Cann et al. paper, there has been very little direct feedback on this subject given the same molecular marker as well as methods. Though this research tends to be of great interest to the anthropological community as well as the general public, very few anthropologists are qualified to perform such analyses. The majority of the studies have been done by geneticists or biochemists, and often the results do not relate to the known elements of the fossil record. Though it is correct that much of the fossil record is subject to interpretation, a great deal of it is well established and there is little room for adjustment to the recent findings based on molecular data. The year 1991 marked the beginning of a great deal of research aimed towards uncovering the mystery that lies within our DNA. Several teams were working at this time to sequence different portions of the genome. While Vigilant et al. concentrated on a single portion of the DNA, Merriwether et al. (1991) focused their attention on several different portions of mitochondrial DNA as well as several different nuclear DNA markers. This study has provided more information on the phylogenetic relationships of the global population than any of the studies published as of yet. In this particular study, mitochondrial DNA of 3,065 humans from 62 geographic areas were analyzed. The data from this research will again be discussed in analysis and methods where it will be used as a standard by which all of the sequencing and phylogenetic tree construction in this paper can be measured. The importance of this study is further amplified by the diversity and sample size of each of the ethnic groups. The African population consisted of 22 ethnic groups spanning the entire continent with a sample size of 863 individuals. The Caucasian population consisted of 15 ethnic groups and a sample size of 1,478 individuals. The Asian population consisted of 18 groups of people and 552 individuals, covering a large area on the Asian continent as well as on the Pacific Rim islands. The last population to be considered consisted of seven Native North American Indian groups, comprising 172 individuals. From these several populations, dendrograms were created to determine the (3 of 5) [11/30/2007 1:20:01 PM]

8 LITERATURE REVIEW relatedness of the populations. Dendrograms are analogous to phylogenetic trees, except comparisons are made within the same species, instead of implementing an outgroup. The dendrogram appears as a forked tree in which the most diverse population is at the top, followed by an interconnection leading to the most homogenous population at the bottom of the dendrogram. From the Merriwether et al. dendrogram (1991), South Africans were determined to have the highest nucleotide divergence. One of several problems with the Cann et al. (1987) paper was the sample size and geographic distribution. It appears as though Merriwether et al. (1991) have rectified this problem by using a geographically diverse population in their sampling. Though this was the most commonly cited problem with the Cann et al. paper (1987), the greatest nucleotide divergence lies in Africa, which is what was originally suggested in David L. Swofford developed the PAUP program, which has a long history of phylogenetic analysis and inference associated with it. It has been revised several times and Version 4.0 (1999) is the most current. It is not yet available on the market but can be obtained for test use in academia. PAUP functions in finding and evaluating phylogenetic trees under the minimum evolution, maximum likelihood, and parsimony criteria. The reliability of these trees may be tested using different methods, among them the permutation tests and minimum likelihood scores. PAUP includes extensive pairwise distance calculations, consensus techniques and reconstruction of ancestral states using parsimony and likelihood methods. Though PAUP is the most commonly cited program for phylogenetic analysis, PHYLIP, authored by J. Felsenstein, is also frequently used. Though PHYLIP has the same capacities as PAUP, it also has additional applications, which allow one to perform compatibility analysis, assessment of independent contrasts and consensus analysis. Proceed (4 of 5) [11/30/2007 1:20:01 PM]

9 LITERATURE REVIEW (5 of 5) [11/30/2007 1:20:01 PM]

10 DATA DATA All analyses of scientific merit must have a reference source from which the experimental data can be compared to. For this reason, the present study consisted of two analyses. The first analysis was carried out by interpreting the dendrogram found in figure 5 of the Merriwether et al. paper (1991), and served as the point of reference. The phylogenetic tree was broken down into degree of nucleotide divergence between populations. Subsequently, these populations, each possessing an assigned number corresponding to degree of nucleotide divergence, were then placed on a geographical map. The amount of nucleotide divergence is a direct measure of age of the population. According to the tree, the Hualapai and Johannesburg populations are the most deeply rooted in the tree, indicating them as the oldest of the populations included in the Merriwether et al. study (1991). The newer populations follow, with the most recent population according to this phylogenetic tree possessing the least amount of nucleotide divergence. These turn out to be the populations of the South African Jews and the Ainu. The second analysis is the actual collection of data for this study. MitBase was initially searched for sequences of mitochondrial DNA from the D-loop region. The available sequences were carefully analyzed, and it was determined that the majority of sequences available on MitBase had a fragment length of 360 base pairs. For this reason, it was decided that a fragment length of that size would be used in the study. It was at this point that a command known as a complex query was executed and the Human MitBase (a division within MitBase) was searched for sequences with a fragment length of 360 base pairs. The search turned up a large set of data, consisting of sequences of over 400 individuals. Approximately 25% of the 400 individuals were used for this study. Appendix 1 contains a sample of the actual data used for this analysis. Each sequence in MitBase is accompanied by several different accession numbers, which can link it to PubMed and Medline; two search engines that can be used to obtain sequence information as well as citations and references. Also included is the reference from which the original data using the sequences was published. Below the reference is the geographic locale of the sample, as well as specific information such as tribe, or ethnic group. Listed at the very bottom is the sequence information, appearing in groups of 10 bases. Other miscellaneous information provided is the molecular source of the mtdna as well as the type of analysis that was undertaken, the most common of which are the PCR/ Sanger sequencing analysis. Whenever DNA sequences are to be analyzed, an alignment must first be carried out. An alignment of the sequences is obtained by inserting the gaps, which correspond to insertions or deletions, into one or more of the sequences in order to place positions inferred to be homologous into the same column of the data matrix (Hillis et al., 1996). It is desirable to have sequences of the same fragment length to make it possible to carry out a more accurate alignment. Based on MitBase availability, a fragment length of 360 base pairs was used. This fragment originated from the region in the mtdna. The program implemented to align the sequences was MacVector, available on Macintosh computers found in the sequencing facility of the Illinois State University Department of Biological Sciences. In order to use MacVector, one must have the sequences for analysis available. These can either be copied from another application and then pasted into MacVector, or they can be input manually. For this analysis, all data was entered manually, as these sequences were obtained from an IBM computer, which is not compatible with the Macintosh computer in the sequencing facility. Each sequence is entered in its own folder, and upon the addition of all desired sequences, the comparisons can be made. After the entry of each of the sequences from the sample, each folder is recalled individually. The sequence is then commanded to "Align to Folder," which generates a relatedness between the other sequences. The first item generated from this command is the description list, a sample of which can be viewed in Appendix 2. The description list consists of a list of the sequence entries, appearing in order from most similar to least similar. The program next generates the horizontal map, which can be viewed in Appendix 3. This visually illustrates differences in the sequences, employing either a slash through the sequence in the area where it occurred (line 14, Appendix 3), or by a white box (line 14, Appendix 4). Again, the sequences are placed in order from most similar to least similar. The final data to be generated when aligning to the folder is the alignment list, Appendix 5. The alignment list provides a visual comparison of the actual sequences entered, with results ranked in order from most similar to least similar. Though this is the most tedious comparison of the three as it encompasses over 40 pages of comparisons, it allows one to inspect the actual locations of the substitutions. These comparisons were made for each sequence, and after the comparison of the first sequence, only the description list was generated, as it gave all of the information of particular relevance to this study. CONTINENT ETHNIC GROUP MOLECULAR IDENTIFICATION SOURCE NUMBER North America Nootka Serum 1 North America Nootka Serum 2 North America Nootka Serum 3 North America Nootka Serum 4 North America Nootka Serum 5 North America Nootka Serum 6 North America Nootka Serum 7 North America Nootka Serum 8 North America Haida Serum 9 North America Bellacolla Serum 10 North America Nootka Serum 11 North America Nootka Serum 13 North America Haida Serum 14 North America Bellacolla Serum 15 Africa/Nigeria Hausa Whole Blood 1721 Africa/Nigeria Fulfulde Whole Blood 1720 Table 1: This table indicates the individuals that were input in MacVecotr. Column one indicates the continent on which the ethnic group was found, followed by column two, the name of the specific ethnic group. Column three shows the source of the mitochondrial DNA, follwoed by the the identification number in column three; which is a reference to MitBase. Africa/Kenya Kikuyu Whole Blood 1738 Africa/ Kenya Kikuyu Whole Blood 1739 Africa/Kenya Kikuyu Whole Blood 1740 Africa/Kenya Kikuyu Whole Blood 1741 Africa/Kenya Kikuyu Whole Blood 1742 Africa/Kenya Kikuyu Whole Blood 1743 Africa/Kenya Kikuyu Whole Blood 1744 Africa/Kenya Kikuyu Whole Blood 1745 Africa/Kenya Kikuyu Whole Blood 1747 Africa/Kenya Kikuyu Whole Blood 1748 Africa/Kenya Kikuyu Whole Blood 1749 Africa/Nigeria Fulfulde Whole Blood 1750 Africa/Nigeria Fulfulde Whole Blood 1751 Africa/Nigeria Fulfulde Whole Blood 1752 Africa/Nigeria Fulfulde Whole Blood 1753 Africa/Ethiopia Somali Whole Blood 1821 Africa/Ethiopia Somali Whole Blood 1822 Africa/Ethiopia Somali Whole Blood 1823 Africa/Ethiopia Somali Whole Blood 1872 Africa/Ethiopia Somali Whole Blood 1873 Africa/Nigeria Fulfulde Whole Blood 1874 Africa/Nigeria Fulfulde Whole Blood 1875 Africa/Nigeria Fulfulde Whole Blood 1876 Africa/Nigeria Fulfulde Whole Blood 1877 Africa/Niger Songhai Whole Blood 1878 Asia/Indonesia Roti Whole Blood 2070 Asia/Indonesia Hiri Whole Blood 2072 Asia/Indonesia Alor Whole Blood (1 of 2) [11/30/2007 1:20:02 PM]

11 DATA Asia/Indonesia Timor Whole Blood 2076 Asia/Indonesia Flores Whole Blood 2078 Asia/Indonesia Roti Whole Blood 2080 Asia/Indonesia Flores Whole Blood 2082 Oceania/Papua New Guinea Papuan Whole Blood 2084 Oceania/American Samoa Samoan Red Blood Cells 2086 Oceania/American Samoa Samoan Red Blood Cells 2088 Oceania/Papua New Guinea Motu Whole Blood 2090 Oceania/American Samoa Samoan Red Blood Cells 2092 Oceania/Papua New Guinea Rabaul Whole Blood 2094 Oceania/Papua New Guinea Sissano Whole Blood 2096 Oceania/American Samoa Samoan Red Blood Cells 2106 Oceania/Papua New Guinea Eleme Whole Blood 2098 Table 1 consists of the continental location of each sequence, the ethnic group, and the molecular source. The identification number is an accession number assigned by MitBase. This number makes it possible to trace back the source to the database if any questions should arise. The database can be found at The actual data generated from this analysis is complex and difficult to put into tabular form. From the MacVector alignments, it is quite apparent that within each population the sequences are more similar than when compared between populations. However, within the African population there was a great deal more diversity in the samples, even within the same ethnic group. Upon comparing sequences it was determined the most homogenous of all were the Oceania individuals. Proceed (2 of 2) [11/30/2007 1:20:02 PM]

12 ANALYSIS ANALYSIS References in MitBase accompany each of the sequences, and indicate the study for which the sequencing was originally performed. Since each sequence had previously been analyzed, it was helpful to refer to each accompanying article to interpret the computed data MitBase generated. The first analysis, the Merriwether et al. Study (1991) exhibits a similar phylogenetic relationship with respect to all populations surveyed. This study was a compilation of several previously published studies that were synthesized to generate a single phylogenetic tree. The Merriwether research is similar to the current study presented in this paper as no original data was collected but instead took advantage of pre-existing information of mitochondrial DNA to correlate to several molecular markers. As has already been pointed out, a study must implement a varied range of molecular markers to obtain accurate results. This is why the Merriwether et al. study and ones like it are considered to be the more complete pictures of human origin currently available in the literature since they are syntheses and averages of many studies. Upon examination of the results obtained from my alignment study, it is quite apparent that, although performed on a much smaller scale, the results do compare favorably with the Merriwether et al. study. Although my data, along with several other's data (Cann et al., 1987; Merriwether et al., 1991 and Vigilant et al, 1991) seem to point to an African origin of modern humans, one must use caution in making such a claim. The data generated from MacVector is subject to interpretation, as there is no hard and fast comparison on all of the sequences together. I however find this program to be more beneficial, however, than PAUP, due to the fact that PAUP is a simulation whereas MacVector is a comparison that one can do visually with all of the sequences available. There are several factors that could potentially be attributed to the intense variation that is exhibited within the African population. First of all, this variation may be attributed to colonization and interbreeding, which would introduce different genetic backgrounds into the population. One must also consider the size and geological formation that exist in Africa in relation to all of the other geographic areas under consideration. Not only is Africa extremely large, allowing for isolation of groups, but many of the population groups are surrounded by deserts, making gene exchange extremely rare and difficult until quite recently. This inhibition of gene exchange can function very similarly to intense gene flow when comparing ethnic groups, as in both cases the ethnic groups can potentially be very different from each other. Proceed (1 of 2) [11/30/2007 1:20:03 PM]

13 ANALYSIS (2 of 2) [11/30/2007 1:20:03 PM]

14 CONCLUSION CONCLUSION At this point, it is necessary to reflect upon what was accomplished in this study. The databases available on the internet were searched for D-Loop regions of mtdna that had published sequences. It was determined that the most commonly cited fragment length was a sequence 360 base pairs long, so that was what was used for this analysis. These sequences were then printed and taken to the sequencing facility in the Illinois State University's Department of Biological Sciences. Each sequence was named according to its geographic location and subsequently typed into the computer program, MacVector. This computer program has the capability to compare all of the sequences to each other to determine relatedness within and among the different populations. According to MacVector, there was more similarity within the Oceania population than within any of the other groups. The African groups, when compared to each other, possessed the most variance; this was in clear agreement with the scientific literature and the largely accepted idea that the variation in African sequences can be interpreted to mean that they possess a longer genetic history as Homo sapiens. The publication of the Cann et al. paper (1987) has raised some very curious questions about the origin of modern humans, and has provided a mode by which it may be possible to further unravel this great mystery. When Vincent Sarich and Allan Wilson initiated the molecular revolution in anthropological applications the power that the genetic code holds was not realized until it was suggested that this data could be construed to mean our family tree is deeply rooted in Africa at a date of around 200,000 years ago. Though this date, initially proposed by Cann et al. (1987) and subsequently by several others, has been widely held within the scientific community for over ten years, some rapid changes have been occurring that point to a much older age. Based upon current research, it is now proposed that mtdna is not strictly inherited from the female line, but may potentially be inherited from males as well. With this proposition, the age of our lineage can almost double. The current research presented here is simply the edge of a vast amount of data and experimentation that is certain to continue with the advent of new techniques. The gathering of additional information will need to be cross-referenced and collaborated. This is sure to help researches determine a more conclusive age and point of origin for the "Mitochondrial Eve". Proceed (1 of 2) [11/30/2007 1:20:03 PM]

15 CONCLUSION (2 of 2) [11/30/2007 1:20:03 PM]

16 REFERENCES CITED REFERENCES CITED Avise, John 1. Molecular Markers, Natural History and Evolution. New York, NY: Chapman and Hall. 2. Ayala, Francisco 3. The Myth of Eve: Molecular Biology and Human Origins. Science 270: Benson, Dennis A. et al GenBank. Nucleic Acids Research 27: Cann, Rebecca, Mark Stoneking and Allan Wilson 1. Mitochondrial DNA and Human Evolution. Nature 325: Hillis, David M., Craig Moritz and Barbara K. Mable Molecular Systematics. Sunderland, MA: Sinauer Associates, Inc. Horai, Satoshi, et al. 1. Recent African Origin of Modern Humans Revealed by Complete Sequences of Hominoid Miochondrial DNAs. Proceeding of the National Academy of Science of the United States of America 92: Jorde, Lynn, Michael Bamshad and Alan Rogers 1. Using Mitochondrial and Nuclear DNA Markers to Reconstruct Human Evolution. BioEssays 20: Krings, Matthias, et al. 1. Neandertal DNA Sequences and the Origin of Modern Humans. Cell 90: Merriwether, D. Andrew et al. 1. The Structure of Human Mitochondrial DNA Variation. Journal of Molecular Evolution 3: (1 of 3) [11/30/2007 1:20:04 PM]

17 REFERENCES CITED Pasarino, Giuseppe, et al. 1. Different Genetic Components in the Ethiopian Population, Identified by mtdna andy-chromosome Polymorphisms. American Journal of Human Genetics 62: Redd, A. J. et al. 1. Evolutionary History of the COII/tRNA lys Intergenic 9 Base Pair Deletion in Human Mitochondrial DNAs from the Pacific. Molecular Biology and Evolution 12: Relethford, John 1. Mitochondrial DNA and Ancient Population Growth. American Journal of Physical Anthropology 105:1-7. Ruvolo, Maryellen 1. Molecular Phylogeny of the Hominoids: Inferences from Multiple Independent 2. DNA Sequence Data Sets. Molecular Biology and Evolution 14(3): Seielstad, Mark T., Eric Minch and L. Luca Cavalli-Sforza 4. Genetic Evidence for a Higher Female Migration Rate in Humans. Nature Genetics 20: Templeton, Alan 1. Human Origins and Analysis of Mitochondrial DNA Sequences. Science 255: The "Eve" Hypothesis: A Genetic Critique and Reanalysis. American Anthropologist 95(1): Vigilant, Linda, et al. 1. African Populations and the Evolution of Human Mitochondrial DNA. Science 253: Ward, R. H., et al. 1. Extensive Mitochondrial Diversity Within a Single Amerindian Tribe. (2 of 3) [11/30/2007 1:20:04 PM]

18 REFERENCES CITED Proceedings of the National Academy of Science 88: Watson, E., et al mtdna Sequence Diversity in Africa. American Journal of Human Genetics 59: (3 of 3) [11/30/2007 1:20:04 PM]