Molecular systematics Spring 2018

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2 Credits- half-semester module, 19 March-29 April 2016

Lectures: M & W 11:00-12:15 Bio-Pharm 3rd floor conference room.

M 2:30-4:30; Th 2:00-4:00 (Each lab session starts in 3rd floor conference room then moves to BioPharm 325).

Chris Simon, Biopharm 305D, 6-4640, <> Graduate Assistant: Katie Taylor, TLS 479,, 6-5479

Readings: will be posted as PDF’s.

Optional reference books: 1) Paul Lewis's unpublished text; 2) The Phylogenetic Handbook (eds. Philippe Lemey, Marco Salemi, and Anne-Mieke Vandamme, 2010); 3) Inferring Phylogenies (Felsenstein 2004, Sinauer); 4) Molecular Evolution: A phylogenetic Approach (Page & Holmes 1998, Blackwell); 5) Molecular Systematics, 2nd ed. (Hillis, Moritz & Mable, eds. 1996, Sinauer) especially Chapter 11 by Swofford et al. on Phylogenetic Inference.

Lecture Goals: The course will focus on the basics of molecular systematics theory and practice from the point of view of the data. We will explore the ways in which an understanding of processes of evolution of molecular data can help in the construction of evolutionary trees. Lectures will examine some of the most serious problems in evolutionary tree construction: nucleotide bias, alignment, homoplasy, among-site rate variation, taxon sampling, long branches, big trees, heterogeneous rates of evolution among branches, covarion shifts.

Laboratory Goals: Labs will cover basic techniques in molecular systematics from DNA extraction to sequencing, alignment and cloning. This lab will be of interest to both experienced and novice molecular systematists because we will try newly developed kits/techniques and compare them to older ones and we will pursue a class project.

Short Assignments:

1) For each topic a bibliography will be provided including one focal paper for which the PDF will be posted. Each student will need to turn in a one-page summary of the importance of each focal paper (1 or occasionally 2 papers per week).

2) The week prior to the start of classes you will be given a checklist discussing practical considerations, organization and data checks for molecular systematics. In certain sections you are asked to answer questions and explain how these procedures are modified in your lab.

3) There will be a short "secondary structure alignment assignment" during the semester.

4) Each student will keep a laboratory notebook and hand-in data collected during the course in the form of an alignment and a nexus data file. Various exercises will be performed in laboratory and some will be finished outside of class. These are detailed in the laboratory syllabus.

5) For each Lab, one student will present a 10-15 minute Powerpoint presentation relating to techniques used in that day’s lab. Ursula will be available to advise you, but use web searches and try to do as much as possible on your own. These Powerpoint presentations will be posted on the class website so that in the future when you teach a molecular systematics class, they can be used as a starting point to revise and develop lectures of your own.

Final Exam: The final exam will be a take home test in which each student critiques the first draft of a paper submitted to Systematic Biology (submitted in the past but making comments as if it were submitted today). Each student will also compare the submitted version to the published version. The answer key will be the actual review containing reviewers, associate editors, and editor’s comments (with permission of authors, reviewers and editors) and a list of critical points that need to be considered by the authors.

Final Due Dates: Sunday 1st May: Lab project and notebook due. Take Home FINAL EXAM handed out. Sunday 8th May: Take home final due.



Day Topics Reading/Assignment/Bibliography Lab
Mar 19
Lecture 1. An introduction to looking at your data: How molecules evolve. Data checks Pdficon small.gif
Read Simon et al. 1994. 651-670 (up to the section that starts on the bottom of the second column). Too large to post, will be emailed to you. How Molecules Evolve & Model Choice Bibliography: Pdficon small.gif LAB: Data checks at every step. Mechanics of Lab; Qiagen kit extractions. Qiagen kit extraction protocol: Pdficon small.gif
Wednesday Mar 21 Lecture 2. How molecules evolve, continued. Pdficon small.gif
Read Sullivan and Swofford 2001 for Monday March 26th. Among Site Rate Variation Readings: Pdficon small.gif Mini-presentation: DNA extraction (Katie) Pdficon small.gif
LAB: Qiagen extractions continued and plant extractions. CTAB plant extraction protocol: Pdficon small.gif
Mar 26
Lecture 3. ASRV, models of evolution, and the history of molecular systematics. Calculating the probability of substitution for sites, Fitch and Margoliash invariant sites models & negative binominal models,Weighting stems and loops. Pdficon small.gif Mini-presentation: Primer Design ( Katie ) Pdficon small.gif
LAB: Explanation of class Tettigades project Pdficon small.gif and Making gels, running extractions on gels, DNA extraction quantification, Troubleshooting and improving “universal” primers for COI.

Qubit protocol: Pdficon small.gif Nanodrop manual Pdficon small.gif Nanodrop protocol: Pdficon small.gif

Wednesday Mar 28 Lecture 4. Correlated changes- should consider stems vs loops; How much to down weight and how to partition when weighting is problematic; Different methods for calculating & accommodating ASRV; For probability of substitution, using a tree is more effective than an alignment; The interaction of tree shape and ASRV; The two components of evolutionary trees; (equal weights aka evenly weighted; misnomer “unweighted” parsimony); Effects of Ignoring ASRV Pdficon small.gif Read for Monday April 2nd, Bull et al. 1993. Classic paper from the Hillis Lab on partitioning and combing data, Bull et al. 1993. Pdficon small.gif Mini-presentation:The polymerase chain reaction ( Zoe ) Pdficon small.gif
LAB: Setting-up PCR reactions. PCR protocol: Pdficon small.gif
Apr 2
Lecture 5. History of “combining data”, As many kinds of data as possible, non-specificity hypothesis, To combine or not to combine? That is the question. Lack of agreement among character subsets, Random error vs systematic error, Assumptions of combined analysis, Bull et al. vs. Chippindale & Wiens; ASRV &ALRV, Homothermia Pdficon small.gif Read and Summarize for Class by Monday April 9th Pagel, M. and A. Meade. 2004. Read and Summarize for Class on Wednesday, April 11th Kainer, D. and R. Lanfear. 2015. Combining Data, Partitioning, Species Trees readings Pdficon small.gif, Pagel and Meade 2004 Pdficon small.gif, Kanier and Landfear 2015. Pdficon small.gif Mini-presentation: Different methods for cleaning PCR products for sequencing reactions ( Tanner ) (Tanner's presentation is lost, but here is a presentation on the same topic from a prior year of the class, which might be a useful reference Pdficon small.gif )
LAB: Running PCR products on gels, purifying PCR products with ExoSAP-IT, and setting-up sequencing reactions. PCR clean up protocol and Cycle sequencing protocol: Pdficon small.gif
Wednesday Apr 4 Lecture 6. Tests for combining data; testing whether the same tree underlies each data partition. Partitioning; Choosing among models for pre-assigned partitions; Automated partition assignment and partition simplification; Model averaging and mixture modelsPdficon small.gif
Mini-presentation: Numts ( Johnny ) Pdficon small.gif
LAB: Cleaning and putting samples on the ABI; Looking at sequences using Geneious. Sephadex cleaning protocol and loading ABI machine protocol: Pdficon small.gif
Apr 09
Lecture 7. What is a long branch?; The meaning of “basal”; Node density artifacts; Felsenstein 1978- when will parsimony be positively misleading?; Penny & Hendy 1989- long branch attraction; Huelsenbeck & Hillis simulations to explore tree space. Accuracy of different phylogenetic methods; Swofford et al. 2001. Bias in Phylogeny estimation due to long branches: Parsimony vs. likelihood in tree space; Remaining uncommitted Pdficon small.gif Covarion, Heterotachy, Nucleotide Bias Readings Pdficon small.gif

Read and summarize for Class (Due Monday, April 16) Gruenheit, Nicole, Peter J. Lockhart, Mike Steel, and William Martin. 2008. Pdficon small.gif
Mini-presentation: How Big Dye works, chromatograms, and troubleshooting(Diler )
LAB: Viewing and interpreting sequencing results, setting up long range PCR. Long range PCR protocol Pdficon small.gif
Wednesday Apr 11 Lecture 8. ALRV: heterotachy, covarion models;Among Lineage rate variation: Covarion evolution: codon models Pdficon small.gif
Mini-presentation: : Depositing sequences in GenBank ( Tanner )
LAB: Running long rage PCR gelPdficon small.gif , cleaning long range PRC product, setting up 2nd short PCR, Make reagents for bead cleanup protocol. Protocol for making bead cleanup mix and testing: Pdficon small.gif
Apr 16
Lecture 9. Heterotachous evolution continued, Covarion Models, The Case for Stationary Genes, Mixture of Branch Lengths for building trees and studying selection. Covarion Mixture Models.Pdficon small.gif Mini-presentation: Ancient DNA & Museum DNA protocols ( Zoe )
LAB: Running short rage PCR gel Pdficon small.gif , cleaning PRC product, setting up sequencing reaction, test bead clean up method Pdficon small.gif
Wednesday Apr 18 Lecture 10: Problems associated with nodal support Pdficon small.gif Nodal Support Readings Pdficon small.gif

Read and Summarize for Next week.... Monday 23 April 18. Salichos L, Stamatakis A, Rokas A. 2014. Novel information theory-based measures for quantifying incongruence among phylogenetic trees. Molecular Biology and Evolution 31:1261-1271.Pdficon small.gif (No need to summarize the derivation, just the introduction and the applications).

Mini-presentation: RNA: extraction and what it can be used for ( Diler )
LAB: Cleaning and putting samples on the ABI, and starting RNA isolation with Trizol. Trizol RNA from tissue protocol: Pdficon small.gif
Apr 23
Lecture 11) Nodal support continued. Spectral analysis, Internode certainty, SplitsTrees. Misc. topics: Big Trees; more taxa or more sequences. LAB: Finish RNA isolation, Compare sequencing results from long range and typical PCR
Wednesday Apr 25 Lecture 12: Secondary structure & alignment. Combining nuclear and organelle data for evolutionary insights on speciation and hybridization
Mini-presentation: Gel electrophoresis ( Johnny )
LAB: Katie presents on Next Generation Sequencing and Applications
Sunday April 29th
Lab notebook due. Take home final handed out.
May 6th
Final Exam due, emailed to Katie who will transmit the anonymous papers to Chris along with a list of pseudonyms

Final Exam Files