More news from the lab and this time its me ranting about the behaviour of my cells.
When investigating the function of a particular gene one tactic geneticists have availible to them is to break the gene and see what happens. One way of achieving this is to ‘knock it out,’ to entirely remove the gene from the genome, or otherwise inactivate it. This allows you to study the phenotype (Characteristics) of the resultant cells to try and work out what the gene was probably doing. However in some cases the gene is very important indeed, and the resultant phenotype is merely ‘dead’ which makes it very difficult to actually do anything with the cells, and impossible to get enough to actually characterise them. Dead isn’t a very usefull phenotype, we want to know exactly what type of dead they are.
One way of overcoming this hurdle is through what is known as a conditional mutant. As the name suggests, this is a mutated version of a gene which only shows its abberent behaviour under certain ‘restrictive’ conditions. One of the most common forms of conditional mutant is the temperature sensitive mutant, which only shows its ‘dead’ phenotype at higher temperatures. Thus most work is conducted at a lower temperature and then the cells are shifted to a higher temperature for testing.
The only problem is that even at the cooler temperature temperature-sensitive mutants are rarely wild-type (normal) and are usually at a slight disadvantage, showing slower growth than normal cells. Now as any student of Darwin will realise we have a bit of a problem. Genomes mutate with a low background frequency which is nethertheless high enough to cause problems, as any mutations that help rescue the slight disadvantages will be selected for, either by outcompetition in liquid culture, or by selection bias (which is basically me selecting the bigest colonies on my plates). Unfortunately sometimes these new mutations also help to make the cells grow better at the higher, normally restrictive temperature, and better than dead happens to be alive.
Now this is very annoying to me, as I want my cells to be dead at the restrictive temperature. For one it identifies where my mutant gene is, and for another is important for other experiements in which I try and make them not dead through other means (largely by making other mutations, or introducing other genes). But seing as I do most my growth at the colder permissive temperature there is no way of telling if my cells have reverted untill I shift them to the higher temperature, at which point they are dead and of no use to me. Fortunately it is possible to replica plate cells, in which a plat of cells is transfered onto velvet, and then ‘copied’ onto two seperate plates, one of which is kept at the restrictive temperature and the other the permissive. This allows me to make sure I choose cells which grow on one plate but not the other.
Unfortunately I am finding that selection is something which must be maintained almost constantly, and even then revertants still seem to slip through the cracks. It seems sometimes you only realise that something has gone wrong when things grow when they shouldn’t have done.
[tags]genetics, pombe, science, phd, mutants[/tags]
Posts Tagged ‘phd’
More news from the lab and this time its me ranting about the behaviour of my cells.
Its all been a bit hectic recently, as its been the transition from my Masters (Which I passed with distinction) to the PhD. However between this I though I’d take a bit of a break, so spent a few weeks at home, catching up with family and friends. Unlike the move to university there are no forced term dates, so trips home have to be seized as they are appropriate. This means its not really co-ordinated with other people, and besides I now have friends for whom home can be anywhere in the world.
Still, the majority of my friends, excluding those in Edinburgh, are located down South. Disturbingly, outside of Edinburgh it is now London that holds the highest concentration of people I know. I suppose its inevitable, it is after all the Nation’s capital and the largest city in the UK. I just wish it wasn’t quite so blatant about it. Meanwhile I’m stuck out on a bit of a limb, but am no longer alone, as a few friends will be moving up to join me in Scotland, albeit not in Edinburgh.
But outside of my gallivanting back home things have also been busy. I began my PhD mid September and the work is underway. At the moment things are still fairly organised, my lab-book is up to date and I know where and what everything is. I’m surprised this level of organisation has lasted this long; if I can keep it up for the next three year it will make the thesis easy enough to write.
In about eight weeks time I will be expected to produce my ten week report, which is to outline the direction of the project. The main role of this is to focus your work, ensure you know where your heading, and to provide scrap paper when the whole plan changes with the results of the first major experiment.
I’m hoping to update soon, discussing my work (Or rather the background to it. I will avoid the unpublished stuff for obvious reasons), the 2006 Nobel Prizes, a visit from Nobel Laureate Aaron Ciechanover and experiences at Biotechnology YES.
I really need to post here more often. All to often I get a vague idea when I’m out and about, only to never post, whether due to laziness, second thoughts, or forgetfulness. Usually a combination of all three. For example, recent ditched blog topics:
- A comment on an article discussing the general perceptions of scientists
- A review contrasting Psychonauts and Prince of Persia: Sands of Time
- A discussion of the importance of preserving media
- Thoughts on the restrictions DRM places on genuine consumers
- Various discussions of scientific concepts and ideas
- An ode to the SanDisk Cruzer Micro
And with the amount of procrastination I’ve been doing in other fields you’d think things like this would flow naturally. Of course it doesn’t help that my readership is probably at zero, but then again who can I blame for not checking out a blog which is updated less than once a month and largely full of the author complaining that he hasn’t updated. It’s exactly the self obsessed inane waffling that give blogs a bad name. (For good examples of blogs please check my blogroll on the left hand side.)
But enough of that. Tomorrow marks the beginning of the smoking ban in Scotland. Of course, it’ll probably take a while before its truly in place, and I’m sure there will be the odd few places that risk the fines, but essentially over the next few weeks the majority of pubs will become smoke free. Now for me this is wonderful news. I don’t smoke and will greatly welcome the fact that a night out will no longer mean coming home smelling of cigarette smoke; Febreeze must hate moves like this. But it will also avoid the irritated eyes and sick feeling that I get in really smoky atmospheres (While some may ascribe the latter to the alcohol it occurs even when I haven’t been drinking.).
In other news I’ve just finished another section of my Masters course, and have already arranged my return to that lab for the PhD proper. This means that for the next few years this place will be full of vague mumblings about Yeast.
I got hacked, a couple of times actually. The first guy modified this page, the second took control a bit more noticeably. I’ve decided to leave this message here, rather than tracking down the original, as a lesson to myself.
Just hope I’ve patched the vulnerability.
Edit: Oh sod it. I decided to track down the original, and have quoted it below. I even corrected a few of the spelling errors.
After a long period of downtime my blog is finally back online, this time running under different software. Unfortunately the old blog died when my webhost went bankrupt, but through the wonders of Google I have again be able to save the content. I’ll probably just chuck the best of it in a static archive page.
For those of you who don’t know a lot has happened since the last time I posted here the main of which is my graduation and subsequent removal to Edinburgh. Here I’m studying in my Masters year of a four year combined PhD/Masters. I have just recently finished my first mini-project, a ten week lab research project in which not only did I get some decent and novel results but I also wasted several thousand pounds in simple mistakes. The biologists among you will find my addition of DNase to a transcription reaction amusing, the rest of you will be lost.
On Friday we had the unit Christmas party, complete with ceilidh, which proved very exhausting. Naturally, for someone tho has only been to one ceilidh before, I was at a complete loss for half the dances, but thankfully I was not alone in this. We all get another crack at it on Burns Night, so expect lots of grumblings about sore limbs around then. This event also signaled the beginning of the Christmas season in terms of events going on and somehow I seem to have been tied in to about four Christmas dinners, two of which have already happened. I also fear further dinners when I get hope, completely destroying my somewhat naive plans to have my overdraft cleared by January. Still, once Christmas is out the way I should get it paid off fairly quickly. I don’t like being in the red, even when it is interest free. Still, as a general trend income is greater than expenditure at the moment so things shouldn’t be be getting any worse.
My new lab project is working with yeast again, although this time is is S.Pombe, S.cervevisiae’s less famous brother. At the moment things are a bit slow as I’m having to get everything up and ready for the experiment proper, which will probably begin after Christmas. The aim is to over-express a particular gene, making the cells sick in the process. Following this I will over express lots of other genes and look for rescue, that is a gene that can make the sick cells healthy again. In doing so I should hopefully discover what the first gene does, as well as possibly identifying some new genes which have previously been uncharacterized. If it works it should be all very exciting and may mean that I get a chance to name a few genes.