PhD thesis - examinators... enough results ?... (Aug/06/2007 )

>ell i'm here with my manuscript preparation...
I have few results for my thesis. Well i try to do a good overview in the bilbiography and it's quite heavy (i need to do 2 because my thesis covers two big fields...) and now i'm facing some very puzzling questions. So i would be pleased if you may give your opinions or insights on hem.

first question is relevant to the thesis itself. I got few results and is this a matter for defending ? basically i dealed with siRNA exp ress ion to ol. I setted it up. I tried to apply it on a target protein (transcription factor). instead of seeing a decrease of transcription, i saw elevation f it...
so these are quite preliminary results, but is it sufficent to defend ?

second question : composition of the jury.
My boss told me to propose at least 2names of persons i wish to be examinators. But i don't know who to ask. I've decided for one person, but the second i don't have clue. i was wondering how is this done in general ? normally supervisors propose persons doenst they?
finally, i plan (fingers crossed) to defend in decembler. Do you think it's bit late to ask ?...

well these are few questions stresssing me at time.
Thanks for any answer.
fred

I once heard a colleague say that it's not the results that determine whether you are ready to submit your thesis and defend it, it's the experiments you have done. You can't be blamed for experiments that don't give you what you expect (we've experienced the same thing in the lab here, do siRNA and see your protein level increase, so I know how frustrating this can be). In some cases, this effect can be prevented by using less siRNA (for instance see Faitar et al., Genomics 2005). So if you can explain all the steps you did to set up the siRNA, and your supervisor agrees, I wouldn't worry too much. When I first started writing, I thought I had almost no results, but then I went through my lab journals and was able to write a nice thesis (at least that's my opinion )
About the members of the jury, I chose the people whom I had collaborated with during my work, supplemented by somebody my boss proposed. I defended in December and contacted the members of the jury in September, so I reckon you still have time (although it may be different in France of course).

good luck!

Hi fred!
Well I still don´t have own experience about thesis defending but I´ll give you my opinions. I think that the most important thing in a thesis is to have a reasonable hypothesis and have done the appropiate experiments to demonstrate it. If you know (and of course I´m sure you know) how to explain the reason of every step you did, then there shouldn´t be a problem. These parts depend on you (and your boss of course) but then results don´t depend on you, they are what they are. If your boss thinks there are enough results to write a thesis, trust him, and at the end, more relaxed, when you read your finished thesis you´ll see that you have more conclusions and a better thesis than you think now. Don´t be afraid, you will do it very well.

About the jury, I´m not completely sure, but I think here most supervisors comment with their students the composition of the jury. They usually are people whom they have worked with and people with whom you have a good relationship, not "scientific enemies"
I think it´s not late to ask them, although it also depends on when you have your thesis completely finished to send to the jury before defending. Here there are minimum periods of time established by University related to that procedures (sometimes too much bureaucracy ).

Cheer up and don´t let nerves take control!

Hi Fred!!
As Pumuki said is very important a good hypothesis it doesnt matter if is true or false at the end (there is a reason for null hypothesis). If you have results to sustein(? )the discusion and some papers to backup the results you should not have trouble. When i wrote my thesis a few years ago I had a similar problem, the results were against what was expect and against what the "gurus" of the theme said, but I had good results to backup all the thesis. So just write the results in a very clear way, wrote a really good discussion with papers that backup your findings. Important don't write just to fill the papers, but write with sense. Is better a short, very clear and well documented thesis than a long blah, blah thesis that is boring.

For the comitee search people that will help you, not that will be an obstacle(?) and slow down your work. Good luck!! and this is the crucial part...so enjoy each not sleeping night, each cup of coffee and all the good people that help, because when leaving the student life for the working life...you will be nostalgic and will wish to be a student again.

this is probably off topic... but, did you see that recent paper about siRNA which causes increases in protein levels?
According to the paper, the position of where the siRNA is attaching to the transcript can either cause an increase or decrease in expression. it was a very nifty paper. i've stuck on the abstract if anyone is interested.

PNAS, 2006, Nov 14th;103(46):17337-17342


Small dsRNAs induce transcriptional activation in human cells.

Li LC, Okino ST, Zhao H, Pookot D, Place RF, Urakami S, Enokida H, Dahiya R.
Department of Urology, Veterans Affairs Medical Center and University of California, San Francisco, CA 94121, USA. longcheng.li@ucsf.edu

Recent studies have shown that small noncoding RNAs, such as microRNAs and siRNAs, regulate gene expression at multiple levels including chromatin architecture, transcription, RNA editing, RNA stability, and translation. Each form of RNA-dependent regulation has been generally found to silence homologous sequences and collectively called RNAi. To further study the regulatory role of small RNAs at the transcriptional level, we designed and synthesized 21-nt dsRNAs targeting selected promoter regions of human genes E-cadherin, p21(WAF1/CIP1) (p21), and VEGF. Surprisingly, transfection of these dsRNAs into human cell lines caused long-lasting and sequence-specific induction of targeted genes. dsRNA mutation studies reveal that the 5' end of the antisense strand, or "seed" sequence, is critical for activity. Mechanistically, the dsRNA-induced gene activation requires the Argonaute 2 (Ago2) protein and is associated with a loss of lysine-9 methylation on histone 3 at dsRNA-target sites. In conclusion, we have identified several dsRNAs that activate gene expression by targeting noncoding regulatory regions in gene promoters. These findings reveal a more diverse role for small RNA molecules in the regulation of gene expression than previously recognized and identify a potential therapeutic use for dsRNA in targeted gene activation.

V

Hi,
I am writing up my thesis, too, so really not an expert. but my friends and PIs said that the point of your PhD thesis is to show that you are able to think. Results are just to help with this, but it should be important that you can show the reasoning to do your exps, led you to the conclusion... Why you do something, what leads you to that, what support your decision... We have a talking session with a PI about writing and defending thesis, and he said that negative results can be good too if you can explain or reason them out, showing what you learn from it logically.

As for the examiners, actually according to my university, the students are not 'supposed' to know who will be the examiners. My boss chose for me and he wrote to the examiners.

thank for inputs.
V you're not off topic, and an other atricle here describes transcription activation by small RNA duplexes...
nature chem bio for persons who are interested (Nature Chemical Biology vol. 3, no. 3, pp. 136-137 (March, 2007))

for the topic, well i've discussed with a friend yesterday and it seemed he hasn't ask the jury yet.
and finally i though about someone... my boss told me : don't you remember ? he was on the jury of the previous student (the relation between this student weren't at best...)

so i've to think to an other people. and i would stay n french speking persons. The fact is that my boss is really not good at persons manager, but is at really god scientific level and he's really brilliant. That's also a point : i dislike the guy but almost love the scientist... i'm curious but still thinking that all his ex colleagues and persons he collaborate with are in such high level i willl appear like a "puppet" to them... and i didn't make collaborations

Hi Fred,

My story wasn't so different...

I went into the study of iron nutrition and photosynthesis in cyanobacteria with the hypothesis that increased iron would allow the little critters to adapt more quickly to changes in light. I came up with methods to measure their photoadapted state, built computer-controlled light banks, learned to grow cultures of happy marine cyanobacteria, constructed computer models of their physiology and gene expression, bought way too much radioactive bicarbonate, and spent months doing microscopy and scintillation counting. In the end, I found that feeding the bugs more iron resulted in slower photoadaptation -- exactly the opposite of my hypothesis. So, I worked out a new hypothesis suggesting that higher iron levels helps them tolerate shifts in light so they aren't physiologically pressured to photoadapt and presented that, along with some proposed experiments to test that idea, as "future research directions".

...and they all signed off on the dissertation.

For a "future direction" proposal, what about suggesting knockdowns of your target protein using an alternative technology? I'd advocate Morpholino oligos, but there are many to choose from. This way you could show that the upregulation you observed is specific to the knockdown technology employed. Of course, you don't want to propose an experiment that is too easy or they will say "do that and then we'll sign..."

In many cases, the defense is run like a hazing. They work on pushing you past what you can answer and see if you crack. The usual course is that they make it tough for you and then they sign off on the paper. Remember to smile and say "Good question, I don't know" if that's the best response.

Best wishes for the end of this adventure and the opening of the next!

- Jon

hey fred,
what i've heard is that depending on how you write your thesis, depends on the way you denfend it.
for instance, if you present your thesis as 5 chapters: 1. litereature review, 2. -4. published papers, 5. Things that didn't get put in the paper.
You'd have less to defend. A collegue did this, but then again, he's a wizard in the lab.
If you don't have 3 papers (which a vast majority of people don't), then it's a matter of defending why you did the experiments you did.
why use siRNA? why not use a ODN? what about a specific inhibitor? what about other sequences? what about the mthods used to get the siRNA into the system? why not use a transient system? why not use a stable system? why would there be an increase? this is transcriptional or translation elevation?
etc (just playing devil's advocat).

as for people... if you are allowed to choose: choose someone who you like. they don't need to be experts in your field. we're all puppets of someone.
i think you are going to be the best judge of your own work. if you think it is worth defending, it is.


V

I agree with Vetticus3.
Choose someone who you like and who is nice... so that this jury will not give you hard time during the defend.

A friend of mine told me that when he was writing up his thesis, his PI did not like some of the intrepretation of the results...and he had to rewrite it.
I hope your PI are willing to accept your explanation of the results.