Phyloinformaticsor

How to analyze LOTS of sequences

Heath BlackmonUniversity of Texas at Arlington

Bioinformatics – Spring 2014

Phyloinformatic workflowRetrieve Sequences• Phylota• Genbank

Align• MAFFT………………

Evaluate Alignment• LAST• Gblocks / Guidance

Infer Phylogeny

Phyloinformatic workflowRetrieve Sequences• Phylota• Genbank

Align• MAFFT………………

Evaluate Alignment• LAST• Gblocks / Guidance

Infer Phylogeny

www.phylota.net

Select and Download Data

• Find a sequence cluster with:> 500 sequences< 2000 base pairs

- Tetrapoda- Teleostei- eudicotyledons- arthropoda

Select and Download Data

• Find a sequence cluster with:> 500 sequences< 2000 base pairs

Download the example file of 18S sequences from the class google drive: 18S.fa

- Tetrapoda- Teleostei- eudicotyledons- arthropoda

Phyloinformatic workflowRetrieve Sequences• Phylota• Genbank

Align• MAFFT………………

Evaluate Alignment• LAST• Gblocks / Guidance

Phyloinformatic workflowRetrieve Sequences• Phylota• Genbank

Align• MAFFT………………

Evaluate Alignment• LAST• Gblocks / Guidance

ProbConsTCofee

Clustal

MuscleKalign

PRRN

DIALIGN-T

MAFFT

Alignment Programs

Clustal Omega

Bali-Phy

DECIPHER

Balance Between Scalability & Accuracy

Method Score CPU time (s)

Consistency based methods

MAFFT 5.662 86.91 6,000

ProbCons 1.10 87.25 43,000

TCofee 2.46 84.56 210,000

Iterative refinement methods

Muscle 3.52 81.67 3,400

PRRN 3.11 82.61 250,000

MAFFT 3.89 82.16 3,600

ClustalW 2.0 76.67 58,000

Progressive methods

Kalign 1.0 80.25 480

MAFFT 5.662 78.63 140

Muscle 3.52 77.63 160

ClustalW 1.83 75.34 2,000

MAFFT

• Align 1,000s of sequences in minutes/hours• Progressive and iterative methods supported• Multiple scoring schemes

• Install locally or run on the CBRC servers

Go ahead and try aligning the 18S.fa file that you downloaded from the class google drive.

Phyloinformatic workflowRetrieve Sequences• Phylota• Genbank

Align• MAFFT………………

Evaluate Alignment• LAST• Gblocks / Guidance

Dot PlotA

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DELETION / INSERTIONA

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INVERSIONA

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INVERSIONA

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Matches between same strand

Matches between opposite strand

Evaluating the 18S alignment

• Look at your dot plots first. What is wrong with the sequences?

• How would you fix/prevent this problem?

Evaluating Sites in an Alignments

• Bootstrapping - Guidance• ID regions with strong support - Gblocks

GBlocks

GBlocks

GBlocks

9 W residues6 I residues8 F residues

Bootstrapping

Bootstrapping

These scores across the bottom scaled between 0 and 1 report the proportion of alignments that agree on the assignment of nucleotides in the original MSA

Try The Data You Downloaded

• Make an alignment• Check the dot plots• Use Gblocks to remove uncertain sites

– How many sites in initial alignment?– How many sites in filtered alignment?– Did you lose any taxa?

• Treat your alignment as a model parameter!

• BaliPhy: Estimates phylogenetic trees across all possible alignments without conditioning on a single alignment being “true”

• Thanks for listening to me!