Tag Archives: protein

Finding the misery molecule? More like imaging a candidate

Depression, like just about every other mental disease, is a strange and mysterious beast. We’ve reached the stage where most people finally understand that it’s really an illness, really a physical problem, not something can be controlled. We know that there are myriad drugs that can help: citalopram, sertraline, paroxetine, fluoxetine…  But we still can’t reach out and touch it. We still can’t predict it. There’s still not a physical test that you can do. An assay that proves that no, this person isn’t just lazy or melancholy or antisocial: they are depressed.

The headlines

Over the weekend, the news outlets got a bit excited at the prospect that scientists had finally discovered “the molecule responsible for causing feelings of depression” (Independent), “the brain’s most miserable molecule” (The Sunday Times) “the little bastard molecule that causes depression” (msn), “The Thing Responsible for Depression” (Jezebel).

And the science

I wish I could say I’m as excited as they are, but of course the science is never as simple as this. The actual paper released is a slog to say the least, and more Physics than Biology, but I’ll give it a go.  Continue reading

Adventures in protein land part 2: Cytosolic protein extraction

So this is going to be more like a lab book entry than anything else, but I’m finally starting to get my head around some of this protein malarkey and figured I should write it down in multiple places!  Continue reading

Useful Resources (for Wheat Genetics Researchers)

There’s a new PhD student in our lab, and I’ve been trying to sort her out with a list of useful tools and websites. I figured that there are probably plenty of other newbies in the field who are fumbling their way around as I did when I started and could benefit from access to my bookmarks bar, so here goes:

When you first start dealing with sequence you may find it useful to know there are tools for converting all sequence types (e.g. FASTA) to RAW format, which is what you’ll need sequence in for lots of alignment softare to use (I genuinely did it by hand a few times before I found that). Also there are obviously tools for reverse transcribing sequence and translating nucleotides into protein sequence.

BLAST (Basic Local Alignment Search Tool) is the ‘search for a gene or a protein’ database that everybody uses across all fields. Use it to find out what an unknown transcript sequence codes for, or how similar it is to the same gene in other species. (Use BLAST-N for nucleotide sequences, BLAST-P for protein to protein, and X for searching for one with the other)

Cereals DB is the wheat-only search engine as maintained by the university of Bristol. Search for 454 genomic sequence (from the University of Liverpool) – e.g. if you’re building a genomic contig to match your cDNA sequence; ESTs – i.e. a summary of where your gene might be expressed or SNPs (although this doesn’t yet tell you which varieties the SNPs are between).

Once you have some sequence, until you get your hands on proper alignment software (this is assuming you’re using <50 sequences – we’re not talking Next Gen data here) like Chromas, DNAman or Sequencher (the latest version does Next Gen but I’m still on 4.10) Clustal W2 is a handy alignment tool. (You’ll need all sequences going in the same direction, so use the reverse transcribe tool above). It works for protein transcripts too.

To find out a bit more about the function of a protein (e.g. what domains it has) use InterPro Scan (which searches other databases like Panther)

For comparative genomics you should be aware of Gramene  which allows you to search genomic data for a bunch of plants together. That links to the European Nucleotide Archive, which covers similar ground to the BLAST database and the PlantsDB databases, hosted at MIPSfor if you’re interesting in things other than wheat.

You can get access to the shotgun sequencing data from the IWGSC consortium for particular chromosomes, although you need to request access and get a password. This is useful for mapping your gene of interest.

Has anybody got others that I’ve missed?

Adventures in protein land

I’m not a protein biologist.

I wouldn’t usually describe myself as a geneticist either, unless trying to clarify what it is that I spend my time doing, in the same way that for years I would describe myself as playing the violin rather than as a violinist, because that seemed to indicate some level of skill that I didn’t feel I yet had.

But I’m definitely not a protein biologist, because I did no protein work as an undergrad, and I didn’t sign up to do a protein based PhD, and there’s only one post doc in the lab who has the first clue about proteins and – while she is possibly the most helpful post doc ever to cross my academic path – there’s only so much help someone can give you when you don’t know what questions to ask.

Yet in spite of this, I appear to be in a situation where doing protein work is inevitable. A few months ago I learned about PAGE gels and Coomassie blue staining and Western blots. Now, I must actually start extracting protein from real live plants, as opposed to my nice-n-easy-burst-like-a-bubble-E. coli. It turns out that this is rather more complicated than I had hoped.  Continue reading

The Ramen Diaries: Part 8 – Protein Extractions

After a week off with flu, there’s just no avoiding it. Today I have to get back in the lab and do my first ever protein extractions from a plant. I have a protocol, that I am assured works, and no reason to suspect it doesn’t. And I have been sat here for 54 minutes doing admin instead of even going to the greenhouse to get my plants. Looks like the stage fright is back. Le sigh….

Idiot’s guide to Western Blots: Part 1

Having not touched them since … June maybe? Perhaps even May … I’m back to doing Western Blots, a technique that I wasn’t amazingly confident with to begin with. I did a dry run with samples that didn’t matter on Friday, knowing that Iw as likely to make tonnes of little mistakes all over the show, forgetting things that once seemed too intuitive to write down. What I need, I thought to myself, is an Idiot’s Guide.

There’s something that bothers me about Idiots’ Guides: They never tell you where something will go wrong. Or what will happen if it does. So here I present the Baking Biologist’s guide to Western blotting: The things that can go wrong, why they will go wrong, and whether to carry on or just scrap the whole thing.

Continue reading

Keep all those plates spinning

Today’s discovery:

Trying to do 10 simultaneous Western blots is… challenging.

Trying to do 10 Western blots simultaneously in the same day as two rehearsals, an hour of undergraduate teaching and a bikini wax might just be beyond me.

On the plus (down?) side, I was in no state to run this morning, so have slightly more energy right now than I usually would at 2pm on a Thursday afternoon. 

I may re-assess this after the sixth antibody wash of 10 separate membranes!