Post on 26-Dec-2015
“Problems” with morphologicaldata…
• Convergence and parallelisms• Reduction and character loss• Phenotypic vs. genotypic differences• Evaluation of homology• Misinterpretation of change or polarity• Limitation on number of characters• Phenotypic plasticity
Always searching for new types of characters…
Is molecular data intrinsically better than morphological data?
Central Dogma
Lipid pigments: chlorophyll lycopenes xanthophylls carotene
Phenolics: flavonols flavones tannins anthocyanins
Iridoid compounds
Alkaloids(N-containing)e.g. nicotine caffeine morphine betalains
Secondary Metabolites
Terpenes
Development of Molecular (Chemical) Systematic Methods
“Chemosystematics”
• Early methods relied on chromatography to separate complex mixtures of secondary metabolites, detect them, and then compare between taxa “spot botanists” – very phenetic
• Better separation and identification methods developed – used pathway stages as cladistic characters - phytochemistry
• Move away from secondary metabolites to proteins• Early protein studies used immunological reactions• Development of improved electrophoretic methods – permitted
direct protein comparisons between taxa• Comparison of seed storage proteins• Development of direct estimates of genetic relationships based on
allele frequency of enzyme variants
Molecular (DNA) Systematics
• Next step was to examine DNA directly through examination and comparison of restriction fragments (RFLP bands)
• Technology evolved to make it feasible to sequence DNA directly
• Initially limited to single genes or non-coding regions
• Now feasible to sequence large numbers of genes or regions or increasingly even whole genomes relatively quickly
Molecular Systematics- Can obtain phylogenetically informative
characters from any genome of the organism- Assumes that genomes accumulate molecular changes by lineage, as morphological characters do- Possibly greater assurance of homology with molecular data (less likely to misinterpret characters) but homoplasy happens!- Principal advantages are the much greater number of molecular characters available & greater comparability across lineages
Three genomes in plant cells
Chloroplast
135,000-160,000 bp
Generally maternallyinherited
(seed parent)
Mitochondrion
200,000-2,500,000 bp
Generallymaternallyinherited (seed parent)
Nucleus
1.1 x 106
to 1.1 x 1011
kilobase pairs
Biparentallyinherited
Selection of DNA region to compare:
• Should be present in all taxa to be compared• Must have some knowledge of the gene or other
genomic region to develop primers, etc.• Evolutionary rate of sequence changes must be
appropriate to the taxonomic level(s) being investigated; “slow” genes versus “fast” genes
• Sequences should be readily alignable• The biology of the gene (or other DNA sequence)
must be understood to assure homology
Genes frequently used for phylogenetic studies of plants:
• Mitochondrial genome – uniparentally (maternally) inherited, but genes evolve very slowly and structural rearrangements happen very frequently, so generally not useful in studying relationships, but there are some exceptions
• Plastid genome – uniparentally (maternally) inherited- rbcL – ribulose-bisphosphate carboxylase large subunit- ndhF – NADH dehydrogenase subunit F- atpB – ATP synthetase subunit B- matK – maturase subunit K- rpl16 intron – ribosomal protein L16 intron
• Nuclear genome – biparentally inherited- ITS region – internal transcribed spacers ITS1 and ITS2- 18S, 26S ribosomal nuclear DNA repeat- adh – alcohol dehydrogenase- many other genes now with next generation sequencing
Plastid Genome
- Circular, derived from endosymbio- sis of cyanobacteria
- Three zones: LSC (large single copy region) SSC (small single copy region) IR (inverted repeats)
- Genes related to photosynthesis and protein synthesis Fig. 14.4
Automated Sequencing
Scanning of gel to detect fluorescently-labeled DNAs; data fed directly to computer.
How do we analyze molecular variation?
- DNA nucleotide sequences (point
mutations) - Structural rearrangements
-insertions and deletions (indels)-inversions
Insertion-Deletion Events
- Can occur as single nucleotide gains or losses or as lengths of 2-many base pairs- Can also be “chunks” of DNA (i.e., losses of introns)
A molecular synapomorphy for Subfamily Cactoideae (Cactaceae) – deletion of the plastid rpoC1 intron…
(Wallace & Cota, Current Genetics, 1995)
ancestral
derived
North American Clades
South American Clades
Pachycereeae
Corryocactus
“Browningieae I”*
“Browningieae II”*
Cereeae
Trichocereeae
Leptocereeae
Hylocereeae
Shared Deletion 2
- 268 bp
trnL intron deletions – Columnar Cacti
(*Tribe Browningieae polyphyletic)
23 kb inversion in all Asteraceae except for members of Tribe Barnadesieae (now Subfamily Barnadesioideae)
Chloroplast DNA Inversion
Comparative DNA Sequencing• Obtain DNA samples from representative organisms (try
to represent morphological diversity) and outgroups• Identify DNA region(s) for comparison• Extract DNA and use PCR to amplify targeted region• Carry out sequencing reactions• Run sequencing procedures (automated)• Align sequences• Use aligned sequences for phylogenetic analysis
(various programs using various algorithms)• Evaluate data in context of taxonomy and morphology
BEP Clade
Stamensreduced to 3;+ 55 mya
AnomochlooideaePharoideaePuelioideae
Bambusoideae(bamboos)
Pooideae(bluegrasses, wheat)
Ehrhartoideae(rices and allies)
Aristidoideae(wiregrasses)
Panicoideae(maize, panicgrasses)
Chloridoideae(love grasses)
Danthonioideae(pampas grasses)
Micrairoideae
Arundinoideae(reeds)
PACMAD Clade
Crepet & Feldman 1991
Genetic Databases
International Nucleotide Sequence Database Collaboration
GenBank: National Institutes of Health (NIH) Genetic Sequence Database
http://www.ncbi.nlm.nih.gov/genbank/
EMBL: European Bioinformatics Institute Nucleotide Sequence Database
DDBJ: DNA Databank of Japan