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Neanderthals Noonan, et al. Sequencing and Analysis of Neanderthal Genomic DNA Green, et al. Analysis of one million base pairs of Neanderthal DNA Kristine Briedis Journal Club 11/21/06 Neanderthals • Most closely related hominid group to modern humans • First recognized 150 years ago in Germany • Existed 500,000 years ago • Lived in Europe and West Asia • Evolved away from humans • Disappeared 30,000 years ago http://www.msnbc.msn.com/id/13154583/ Interbreeding? • Open question if humans and Neanderthals cohabitated • Overlap in their geographical ranges and time of existence • Some late Neanderthal groups adopted cultural traits that might have been influenced by modern humans DNA • Before now, Neanderthal DNA studies were limited to mitochondrial DNA sequencing (1997) • Sequencing of nuclear Neanderthal DNA would help human genome annotation and identify our most recent genetic differences • Two groups used a bone found in 1980 in a cave in Croatia • Carbon dated to 38,310 ± 2,130 years ago http://www.nature.com/nature/journal/v444/n7117/pdf/444254a.pdf Contamination • Challenge to find uncontaminated samples • Tested >70 Neanderthal bone and tooth samples • Most samples were too degraded or contaminated • Six bones were further tested • Amplified mtDNA that previous studies have shown to be different from modern humans • Vi-80 bone (from Croatia) was best prospect for sequencing Sequencing Approaches • 454 sequencing: • “sequencing by synthesis” • Uses small fragments of DNA (advantageous) • PCR every fragment to 10m copies • Attach DNA to bead • Solutions of nucleotides are progressively applied; photons are generated as each base is incorporated • >1 million base pairs • Metagenomic approach: • Insert DNA fragments into plasmids • Amplify in bacteria • Sequence inserts using either pyrosequencing or Sanger method • Can be used to target specific genes • 65,000 base pairs Vi-80 bone • Recovered 254,933 unique sequences from Vi-80 bone (Nature group) • Aligned sequences to complete genomes, environmental samples (env database), and GenBank set of redundant nucelotide sequences (nt) Neanderthal chromosomes Time of Divergence • Science group: – used SNPs to approximate when modern humans and Neanderthals diverged – Estimated 120,000-670,000 years, with a best guess of 370,000 • Nature group: – used PCR confirmed substitutions of mtDNA – assumed human-chimp divergence 4.7-8.4 millions years – estimate human-Neanderthal divergence of 461,000825,000 years; best guess of 516,000 when including nuclear DNA substitution information Log-likelihood graphs Nucleotide changes • Generated 3-way alignments between Neanderthal, human, and chimp • Estimate 7.1%-8.2% of sequence changes between human and chimp occurred after Neanderthal divergence SNP study • Compared Neanderthal sequences to human SNP sites • Neanderthal has derived (human SNP) allele (as opposed to ancestral chimp allele) in 30% of all SNPs • Possibly overestimation since SNPs analyzed are of high frequency in human and thus are likely older • Estimate conflicts with their population split model • Suggests interbreeding? • Neanderthal X chromosome shows higher level of divergence than autosomes • Possible gene flow from modern human males into Neanderthals Ancestral Population Size • Human effective population size is around 10,000 • Chimps, gorillas, and orangutans have an effective population size 2-4 times larger • Effective population size of human and chimp ancestor thought to be closer to apes than modern humans • Estimate for Neanderthal is up to 12,000 with a best guess of 3,000 • Indicates effective population size of hominids had decreased prior to human-Neanderthal split • Dependent on what split time you use (time after which there was no more interbreeeding between groups) Summary • Papers describe sequencing of about .03% of Neanderthal autosomal genes • Groups agree that human and Neanderthal genomes are <0.5% different • Broad agreement on divergence time (370,000 vs 516,000) • Possible gene flow between human and Neanderthal, but not definitive yet Future Plans • Is it possible to fully sequence genome? • 1x coverage requires 20g of bone and 6000 runs on the 454 sequencing platform • Nature authors think they can make the sequencing 10x more efficient • Their goal is to fully sequence the Neanderthal genome within two years • Currently 5 million bp are available • Science authors are targeting FOXP2 gene – Linked to speech and language; Present in many animals, but human version differs from chimp Happy Thanksgiving! 1-800-BUTTERBALL