Genomics Practical 2008

EST Sequencing: Write Up

This practical will be assessed, and thus you should check the requirements for the writeup
before you continue through the sequence analysis work

The writeup should be handed in to the relevant BOX by 4 pm on Thursday 6th March 2008.

Please "name" your writeup with your matriculation number as this allows us to trace it simply!

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The Writeup, 2008

In your writeup, which should not need to be longer than 6 printed pages, you should

1 In a few paragraphs (~1 page) describe the process by which you isolated the EST sequences.

 Note that this should include comments on why things might have failed (if you had failures).

 

2 Draw up a "molecular CV" (total of ~2 pages) giving the information below for each of the four sequences you choose to analyse. Remember to include quality data supporting each of your annotation statements.

Clone name
Insert length
Sequence length acquired and comments on sequence quality
Position of start of putative open reading frame or initiation methionine; length of open reading frame
Position of polyA tail (if present)
Best BLAST match in nonredundant database
Best BLAST match in mouse proteins
Best BLAST match in arthropod proteins
Domain and protein family membership (if any)
Cellular location inferred, including presence of signal peptide and transmembrane regions, if any

AND, FINALLY: Your putative functional annotation for the gene (in less than 30 words)

3 Choose one of the sequences that had a significant similarity in BLAST searches to proteins from several different organisms, and that you think might play some role in the host-parasite interaction.

a In a few paragraphs, describe the quality of the similarities identified, what the proteins matched in the database do in terms of biology or enzymology in the organisms from which they come, and what you think your gene might be doing in Heligmosomoides polygyrus.

b What role is it likely that your chosen gene plays in the host-parasite interaction between H. polygyrus and the mouse host? Remember to back up any statement you make with evidence from your bioinformatic analyses.

c What experiments could you suggest that would test the role that you have suggested your chosen gene plays in the host-parasite interaction?

 (~2 pages)

4 Answer the following questions (~1 page).

a Why is an EST strategy useful for "neglected" organisms such as the Heligmosomoides polygyrus ?

b Why is an EST strategy also useful for model organisms such as humans and C. elegans where the whole genome has been sequenced?

c Based on what we know about other nematodes, we predict that Heligmosomoides polygyrus will have ~18000 protein coding genes in its genome. If we want to identify as many Heligmosomoides polygyrus genes as we can, how many ESTs should we aim to sequence from how many cDNA libraries? Would sequencing the whole genome (~80 Mb) be more efficient?

 

 

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The writeup should be handed in to the relevant BOX by 4 pm on Thursday 6th March 2008.

Please "name" your writeup with your matriculation number as this allows us to trace it simply!

Mark Blaxter and colleagues for Genomes and Genomics 3, 2008