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Coming into the Genome Age Part II: Exploring Human Variation and Evolution

David Micklos
DNA Learning Center (DNALC), Cold Spring Harbor Laboratory

Updated March 2017

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Why the mitochondrial genome?

In “Coming into the Genome Age Part I,” I challenge biology educators to help students step into the genome age by analyzing DNA with a combination of in vitro lab work and in silico computer work. Here I introduce an experiment that allows students to use their own DNA differences (polymorphisms) to explore human variation, relatedness, and evolution. This experiment examines single nucleotide polymorphisms (SNPs) in the nucleotide sequence of a portion of the mitochondrial (mt) chromosome.

The mt chromosome is an oddity in the human genome. It retains several features of the bacterial chromosome from which it evolved: it is a small, circular molecule (think plasmid) and its 37 genes lack any introns. Because it is located within the mitochondrion, the major source of oxygen free radicals, the noncoding portion of the mt chromosome accumulates mutations at about 10 times the rate of nuclear chromosomes. At conception, sperm mitochondria are excluded and the dividing zygote receives its mt genome only from the cytoplasm of the fertilized egg. Thus, the mt genome is inherited in a maternal lineage. Because there are several copies of the mitochondrial chromosome in each of the hundreds to thousands of mitochondria per cell, mtDNA sequences are highly enriched over nuclear sequences. This increases the chances of retrieving mtDNA from degraded samples from ancient specimens or poorly preserved forensic evidence.

The experiment

The experiment begins with students isolating DNA from cheek cells or hair follicles, then amplifying a 440-nucleotide sequence of the mt hypervariable region by polymerase chain reaction. After checking for proper amplification on an agarose gel, the student amplicons (PCR products) are sent by overnight mail to a sequencing service (small fee applies per sample). Typically in 2-5 days time, the completed sequences are uploaded to the Sequence Server which has an easy-to-use bioinformatics tool to compare 2 or more sequences.

Multiple sequence alignment clearly shows each nucleotide position at which one or more compared sequences differ. These differences are termed single nucleotide polymorphisms (SNPs). Based on the premise that mutations accumulate over time as 2 lineages diverge, a student counts SNPs to gauge her genetic affinity with her classmates. It takes no prodding, and students invariably set up their first experiments to compare their sequences to those of their friends. However, questions mount when subsequent comparisons frequently show a closer relationship (fewer SNPs) with students of different racial or ethnic backgrounds.

Our maternal lineage

Then, students align samples from world populations to gauge the extent of variation that has accumulated in our maternal lineage since modern humans evolved about 150,000 years ago. By pooling results, students can easily determine that any 2 living humans have, on average, 6 to 8 differences over the 440 nucleotides under study. Comparisons between modern humans and ancient Neanderthal samples (29,000 to 100,0000 years old) show approximately 26 to 28 differences, putting Neanderthal outside the variation of modern humans. If 6 to 8 differences equal the 150,000 years since humans diverged from a common ancestor, then 26 to 28 differences would put the human-Neanderthal divergence back 4 times more distant, or about 600,000 years ago.

On the other hand, comparisons show that the 5,000-year-old sample from Ötzi the Tyrolean Ice Man falls within the variation of modern humans. (Typically, in a class of European ancestry, one student will have zero differences!) This raises interesting questions about how different our physiology might be from Ötzi and our hunter-gatherer ancestors, and what this might mean with respect to our "modern" lifestyle.

Out of Africa

Results of 3-way alignments between 2 Africans and one European or Asian again confound commonsense expectations, but provide DNA evidence that overwhelmingly supports a recent ("out of") African origin of all modern humans. At each SNP position, 2 people will have the same SNP, and one will be different. Africans differ at the majority of SNP positions, indicating that African populations have greater genetic diversity. This provides evidence that African populations are older and have maintained relatively larger numbers over the history of modern humans. The non-African matches one of the Africans at the majority of SNP positions, indicating that most European and Asian diversity is a subset of African diversity. This provides evidence that founding European and Asian populations took only a portion of African variation when they migrated out of Africa and that relatively few European- or Asian-specific SNPs have accumulated in the brief time since they split from their African forebears. Thus, this experiment provides a fantastic opportunity for students to discover evidence of human evolution that is written in our own DNA.