Download Multiple Sequence Alignments

Multiple Sequence
Alignments
Assemble DNA sequences into
a ‘contig’
Identify conserved residues
and domains
Multiple Sequence Alignment
of Protein
Sc: yeast
Hs: human
Dm: fly
Ce: nematode
At: plant
Contig Assembly
ABI Sequencing: Relies on
Primer-Directed DNA Synthesis
Chain Terminators are
dideoxy NTP’s
H
ABI Sequencing: Relies on
Primer-Directed DNA Synthesis
ABI Sequencing
ABI Sequencing
• Sequence reads are usually 600-900
bp in length
• Quality of read is poor at beginning
and end
• Quality is best in the middle of the
read
Beginning of an ABI read
Beginning of an ABI read
Middle of an ABI read
Middle of an ABI read
End of an ABI Read
End of an ABI Read
Steps for Contig Assembly
•
•
•
•
Collect ABI files and assess quality
Trim away ends
Compile into fasta format in 1 file
Assemble contig with ‘CAP’ (Contig
Assembly Program)
• Evaluate output - more trimming if needed
• Repeat CAP assembly if needed
• Compare contig with WT or individual reads
and make nucleotide assessments
Protein MSA
• Assemble sequences in fasta format
in 1 file
• Prepare multiple sequence alignment
(MSA) with ClustalW
• Shade conserved residues using
BoxShade
Assemble sequences in fasta format in 1 file
Prepare multiple sequence alignment (MSA)
with ClustalW
Shade conserved residues using BoxShade
Protein MSA
• Modify BoxShade Output for use
– in MS Word doc
– in PowerPoint presentation
– in web page
Modify BoxShade Output in MS Word
In Class MSA Tutorial
• Assemble sequences into a contig
using CAP
• Create a MSA of protein sequences
for use in PowerPoint