CHARACTERS USED IN RECONSTRUCTING PHYLOGENETIC TREES

1. CHARACTERS USED IN RECONSTRUCTING PHYLOGENETIC TREES 1. Morphological 2. Protein analysis Allozymes - different enzyme forms encoded by different alleles at a single locus Electrophoretic profile of allozymic forms of six esterases from six individuals in Drosophila virilis population Forms 2 & 4 show slow (S) and fast (F) migration Avers Fig. 6.16

2. 3. DNA analysis (i) RFLPs (restriction fragment length polymorphisms) M Mitochondrial DNA from 18 different eels cleaved with EcoRI M = size marker Restriction maps of primate nuclear rRNA spacer region Avise,JC Fig. 3.8 (iii) AFLP (amplified-fragment length polymorphisms) PCR based methods

3. (ii) DNA sequencing – align sequences and score nt substitutions How to choose which type of DNA sequence to use? If comparing very divergent organisms: - need ones “universally” present and highly conserved otherwise possible errors in alignment, multiple sub… eg. ribosomal RNAs translation elongation factors, ribosomal proteins glycolytic pathway enzymes

4. Nature of nt substitutions in structural RNA genes - different functional constraints than for protein genes RNA folding – 2o and 3o structure, interactions with proteins in ribosome… - conservation of helical structure important (but not nt sequence in some cases) C - G A - U U - A G - C C - G G - C U - A G - C E.coli 16S rRNA compensatory base changes Brown Fig.11.11

5. If comparing very closely related organisms: need rapidly evolving sequences mitochondrial genes in animals nuclear pseudogenes Human mitochondrial genome Red – protein & rRNA genes Blue – tRNA genes Lewin Fig. 25.5

6. Advantages of using animal mitochondrial sequences High rate nt substitution Maternally inherited, no recombination Easy to isolate & assay - many identical copies per cell Small, well-characterized genome, no repetitive DNA Different regions evolve at different rates ribosomal RNA relatively slow respiratory genes replication origin (D-loop) relatively fast Lewin Fig. 25.5 Biodiversity survey – mt DNA (COI) barcodes (P. Hebert)

7. Dusky seaside sparrow Fig. 5.37 1966 - A.m. nigrescens subspecies on Atlantic coast of Florida endangered species 1980 - only 6 individuals (males) left - artificial breeding program - chose A.m. peninsulae (female) on Gulf coast based on morphological & behavioral characters

8. 1987 – last purebred sparrow died 1989 – mitochondrial DNA restriction analysis for 39 individuals, 11 subspecies Fig. 5.38 - Atlantic coast ones more similar to each other than to any Gulf Coast subspecies - estimated divergence time consistent with change in sea level creating Florida peninsula ~ 0.5 Mya – reproductive barrier

9. PHYLOGENETIC RELATIONSHIPS OF HUMANS AND APES 1950’s - morphological data Human – gorilla split believed to be ~ 20 Mya … and chimp – gorilla split much later Moore Fig. 37.8

10. 1960’s – 1980’s Protein & early molecular data Freeman Fig. 14.2 Human-gorilla divergence time ~ 5 Mya Human – gorilla – chimp trichotomy

11. 1990’s – molecular data Mitochondrial DNA sequences “statistically, this tree is about 10 23 times as likely as the alternative tree…” Y chromosome gene autosomal pseudogenes & nuclear non-coding Human – chimp split more recent than gorilla split Freeman Fig. 14.4

12. Locke, Nature 469:529, 2011 Mitochondrial COII gene Freeman Fig. 14.6 Maximum parsimony tree Branch lengths = minimum number of inferred changes - haplotypes cluster together by species

13. Are sequences from different primates evolving at same rate? 23 Kij = # sub per 100 sites between species i and j **difference significant at 1% level