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Molecular MethodsMolecular Methods
Next: Spring 2016  
Seminar/Exercise for the MSc Module "Plant Population Genetics" (MSc Biodiv&Ecol: A17; MSc LaÖk: V29; MSc MobiPhys FO10)  
V00_Schedule2016 schedule of the seminar 
Seminar 01 – Introduction / how to work with Excel  
V01_Excel_20121021 A short introduction into some useful commands in Excel 
E01_Excel.xls A small Excel file applying these commands 
V01_Intro_20121021.pdf Introduction into the topics of the seminar 
V01=V12_MolMethods_20121021.pdf From DNA extraction to PCR - overview about basic molecular methods 
paper1 manual isolation of DNA 
paper2 manual isolation of DNA 
paper3 manual isolation of DNA 
paper4 manual isolation of DNA 
paper5 manual isolation of DNA 
kit1 Invitek Plant kit for DNA isolation 
Part 1 – Fingerprinting techniques  
Seminar 02 – Basics of fingerprinting, how does AFLP work  
T02_StudTalkA_SexAsex.doc guideline student talk A: Mutation rates, sexual and asexual reproduction in organisms 
xxx Student Talk A  
T02_StudTalkB_ApplicationsGenotyping.doc guideline student talk B: Genotyping and its applications in forensics and evolutionary biology 
xxx Student Talk B  
V02_Intro_Fingerprinting_20121021.pdf Introduction into fingerprinting techniques 
V02_AFLP_principles_20121018.pdf The principles of the AFLP technique 
E02_AFLP_calculator.xls Simulation of the AFLP method: how many fragments will be created and detected in dependence of genome size, adaptor length and fluorescent markers? 
Vos_etal_1995.pdf The paper where the AFLP method was invented 
Seminar 03 – Evaluation of AFLP data  
T03_StudTalkA_RAPD_AFLP.doc guideline student talk A: Pros and Cons of RAPD and AFLP 
xxx Student Talk A  
T03_StudTalkB_Example_AFLP.doc guideline student talk B: Example for application of AFLP in research 
xxx Student Talk B  
V03_AFLP_Gagea_20121018.pdf Example: Application of AFLP to assess the relations of sexual and asexual reproduction in Gagea lutea and G. spathacea 
PSE2011_Gagea AFLP.pdf Research paper: Application of AFLP to assess the relations of sexual and asexual reproduction in Gagea lutea and G. spathacea 
E03_inputAFLP_GAGlutAll_II_AGGCTA.pdf Evaluation of AFLP data with Excel: a typical input file with a 0/1 matrix 
E03_AFLP_GAGlutEH_II_AGGCTA.pdf Evaluation of AFLP data with Excel: simple version without cluster linkage (no chain algorithm) 
E03_AFLP_GAGlutEH_II&III_new.pdf Evaluation of AFLP data with Excel: advanced version with cluster linkage (with chain algorithm) 
E03_SSR_MERper_sim2.pdf  Evaluation of AFLP data with Excel: a small file explaining how cluster linkage works (the chain algorithm programmed, this is highly theoretical!) 
Seminar 04 – Experimental and data evaluation errors with AFLP / software example: Gene Mapper  
E04_AFLP_exercise exercise for creating a 0/1 matrix from AFLP profiles 
S04_AFLP_troobleshooting.pdf Possible experimental errors with AFLP 
BeckmanCoulterAFLP_guide.pdf Manufacturers guideline für AFLP 
Optimization_AFLP_largegenome.pdf original paper: Optimisation of AFLP for plants with large genomes 
Optimization_AFLP_multiplex.pdf original paper: Optimisation of AFLP via multiplexing of different primer combinations 
xxx Link to download the free demo version of Peak finder (a smaller version of GenMapper, does not allow to store and print profiles) 
Seminar 05 – Introduction into microsatellite (SSR) analysis  
T05_StudTalkA_SSR.doc  guideline student talk A: Principles of SSR, comparison with AFLP 
xxx Student Talk A  
http://technelysium.com.au/?page_id=13 Link to download Chromas Lite – a little freeware program to visualise ABI sequencer files 
Seq_SSR_Sphagnum_AK1_A01_2012-12-19_1.ab1 A sequence of a haploid gametophyte of Sphagnum spp., showing a microsatellite locus (motif CT, open file with Chromas Lite) 
S05_SSR_method1 The principles of microsatellite analysis, part I 
S05_SSR_method2 The principles of microsatellite analysis, part II 
Seminar 06 – Analysis of microsatellite data  
E06_Microsatellites_Ing5.xls Example: analysis of microsatellite data with Excel to assess clonality in a Chinese species of Poplar 
TREE_POPeup_Wiehle_etal2009.pdf Example: research paper using SSR analysis to assign roots suckers to adult trees in Populus euphratica 
S06_SSR_data Workflow for SSR data evaluation 
Seminar 07 – troubleshooting of microsatellite data  
T07_StudTalkA_guide_errors.doc  guideline student talk A: Sources for genotyping errors 
xxx Student Talk A  
Microsatellite_errors_HoffmanAmos2005.pdf paper explaining possible sources of genotyping errors 
S07_SSR_errors_experimental How to avoid experimental errors in microsatellite assay 
V07_SSR_errors.pdf Consequences of errors in the evaluation of microsatellite data for the estimation of clonality 
EvolEcol_POPeup_MicrosatErrors2010.pdf Research paper: simulation experiment for consequences of errors in the evaluation of microsatellite data for the estimation of clonality, done for a data set from Populus euphratica 
Part 2 – DNA Sequencing and phylogenetic analyses  
Seminar 08 DNA sequencing and genes  
t08_DNAMethods_2012_introduction.doc guideline for student talks 1-3 of this seminar 
xxx Student talk 1 Principles of Sanger sequencing and PCR 
xxx Student talk 2 DNA sequences for molecular identification (DNA Barcoding) 
xxx Student talk 3 How to use DNA sequences to construct molecular phylogenies 
t08_DNAMethods_2012_talk_1.zip contains 2 papers about DNA Barcoding (material for student talk 2)
t08_DNAMethods_2012_talk_2.pdf Presents a fundamental paper dealing with the importance of genes for phylogeny 
t08_DNAMethods_2012_talk_3.pdf  a paper describing useful genes for molecular phylogenetic reconstruction in fungi 
t08_DNAMethods_2012_working_examples.zip examples for sequencing files (type ab1, to open with Chromas Lite) from the fungal ITS region. 
V08_SequencerABI310_20121018.pdf How does a capillary sequencer work? Short explanation of the components of the device 
Seminar 09 – edit and align DNA sequences  
http://technelysium.com.au/?page_id=13 Link to Chromas Lite, a very short, handy program to view and annotate sequences from the files given by a capillary sequencer (type *.ab1) 
http://www.mbio.ncsu.edu/bioedit/bioedit.html Link to Bioedit, a program to align sequences per hand, annotate them and export into other formats 
http://www.megasoftware.net/ Link to MEGA, another program to align sequences, with a lot of useful features for phylogenetic analyses 
LAM_24701_F_AKlahr_20110429.ab1 example of a raw SSU sequence from a dark spored myxomycete (Meriderma carestiae, *.ab1 file, open with Chromas Lite or MEGA) 
LAM_24701_F_AKlahr_20110429_edit.ab1 edited SSU sequence from a dark spored myxomycete (Meriderma carestiae, *.ab1 file, open with Chromas Lite or MEGA) 
Alignment_exercise1.zip exercise to create and check your own alignment with 10 sequences from the partial SSU of the myxomycete genus Meriderma 
exercise_20131012_MER_EF1a.zip advanced exercise to decipher a heterozygous sequence of the partial EF1a of Meriderma spp. from F and R sequences (zip-File, please download and unpack) 
solution_20131012_MER_EF1a.zip solution of the exercise showing the two alleles in the alignment (zip-File, please download and unpack) 
Seminar 10 – Searching and submitting sequences to data bases (Genbank, EMBL)  
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Seminar 11 – Phylogenetic analyses  
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Seminar 12 – Algorithms to construct phylogenetic trees  
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Seminar 13 – Make your own tree, exercise  
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