hobglobin

Dear JohnSky,

To rule out inhibitors which might exist in your sample, you might consider to spike (or mix, as you call it)  some working RAPD DNA in your RAPD PCR which is not sea cucumber origin (might be from plant, other organism) which is around 10%-20% of the total DNA concentration you had used in your PCR reaction.

I suggest you to do the following:
Saying if your total DNA concentration in for a successful run of "Plant" RAPD PCR is from 500 - 5ng/ul (random figure), in your "sea cucumber" RAPD PCR, try use a total genomic concentration of 100ng/ul in first tube, second tube you mix the PLANT DNA with SEA CUCUMBER in 1:9 ratio (so, for a 100ng genomic content 10ng is from PLANT), third tube just 10ng/ul PLANT DNA, fourth tube SEA CUCUMBER 90ng/ul. Run all your PCR together with your PCR profile. If you only got the third tube working, this means your inhibitor present. IF you got the third and second tube working and failed on the fourth tube, highly possible your RAPD primers is not working. IF your fourth tube is working, this might sounds that you need some dilutions in your template.

Just my 2 cents.

What I do not really understand is the fact that you (the "student") are the one that is "finding" a good PhD topic.

Is this normal in your country?

In belgium its a little bit the other way around: professors find intersting topics and search for PhD people...
Its rare that students come up with their own ideas.. basically because students often arent realistic enough or dont know a lot or not enough to come up with ideas.

Anyway, it seems that you are going to do something with tropical animal husbandry? Why not check research facilities (in other countries) in this area and see what kind of topics they offer?
In belgium for example we have some open topics (still looking for students) , maybe you can get some inspiration from those?
If you want, I can give you some links and perhaps that could give you some inspiration or who know, perhaps you would even like one of them and come the belgium haha

From my experience and comments my supervisor made whilst I was writing up. If you are working exclusively on writing and have strong writing and editing skills then 3-4 months is possible. For anyone with average writing skills at least 6 months writing will be required. Also, you will need to factor into your timing how long it takes your supervisor(s) to return draft's and if they have any upcoming leave or work commitments that may add delays.

As for recommendations on how to write:
Don't try to write the perfect document/chapter the first time. It is more important to just get thoughts and ideas onto the page. Once you have a few pages of notes/comments you will feel a lot more motivated to write than staring at a blank page.

Do your referencing as you go, even if you are just putting notes on the page. It will save you heaps of time in the later stages when you are editing. Also, use a reference manager (EndNote, ProCite, etc.)

Start writing NOW even if you experiments are not finished you can write 1/2 to 3/4 of a chapter without results and then just add the rest later.

And finally when the crushing wave of panic comes (and it happens to everyone) just breathe, step away from the computer and listen to your favourite song/ go for a walk (anything to distract you from your thesis for a few minutes, no longer than 1 hour) and then sit back down and start again.  You will get there in the end.

Good luck!

K.B., on 14 April 2012 - 03:13 AM, said:

I do not see what it is wrong with that question.

Deionized water (demineralized water) is not bacteria free... Its not sterile water...
Same goes for dd water.
The changes to get bacteria in it are even higher since people use the same containers over and over and they are not sterile at all!
Even MQ water can contain bacteria.

I do not know from what country you are, but in my country, tap water is pretty clean.

So the question remains: why boil the water the tap water for the 70% ethanol solution and not, for example, boil the di water?

Also: if you boil the water: the dead bacteria will stay in it.. so you dont remove it, so dont see the point trof made since in di water there are bacteria present too.

PS. the standards for tap water (CFU) in many countries is more strict then those for purified water (often no regulation to CFUs at all)

There is most likely a reason why not to use tap water, but your explenation isnt 100% accurate.

So, not sure whether your *facepalm* should be repeated....


(I am strictly speaking of cleaing a bench, not speaking about using the water for other purposes, ddwater etc has its purpose, but not sure why you need to use it for a 70% ethanol solution.. It has to do with the non bacterial susbtances in the water I think, rather then the bacterial ones)

heroine addict

haha... good one cas... oh well, i know my supervisor has the best intentions...

here's your friday song

timbits

yeah, then her name is going to be hyper long that was my band indeed... had i played that one for you before? what about this one then?

jangajarn, on 21 May 2009 - 05:09 PM, said:

Dear Rangajarn,


The most important thing is LISTEN!!!!!!!!!!!!!!!!!!!!!!!!

Then you can start learning

It does help to have people around you who are worth listening to. Be flexible in what you can do practically. There is a process that occurs when doing experiments in science:-
Read about the area you are going to be experimenting in

Try and meet people who are doing/have done similar work....this reduces the time that can sometimes be lost optimising conditions.....its called the "tricks of the trade" which are not to be found in the "Materials and Methods" sections of published papers.

Prepare thoroughly...have I got all the chemicals, consumables, equipment, SOP's that I require.

Do I have the correct supervision by senior staff when I first start. Do I know how to use all the equipment I need to do the experiment. IS THE EQUIPMENT CALIBRATED SO I CAN DO ACCURATE AND PRECISE AND REPRODUCIBLE EXPERIMENTS.

CONCENTRATE ALL YOUR EFFORTS ON 1 THING....... do not be persuaded to "multitask" i.e. give time and effort to get reproducible and accurate experiments that can be repeated, not only by you but by others!!!!!  Once experience THEN multitask....but beware even experienced researchers make errors if put under too much pressure from the PI/Professors.

I personally found the process daunting when I first worked in a research lab in 1983. However over time you will go through the process many times, and like anything in life the more you do it, the easier it becomes.

Hope this gives you some ideas.

Kindest regards

Uncle Rhombus

First - it depends on the lab you are in - and from what you tell us your lab has not a great reputation in educating people well - because more than one PhD who quits each year is quite a high number.

Most people whom I know finished their PhD within 3-4 years here in Austria. So the time needed sounds relatively long for me - which probably is also not the best sign. Do the people get payed during the whole time of the PhD - usually PhD contracts are given for three years only and then do not get full salary....are you aware of such problems?

High Impact publications are important, but are usually not the things that decide if you get a Post Doc or not - what is more decisive is the amount of contacts you were able to establish during your PhD and what those people think of you - because I know about many many jobs that were never published but were given directly to somebody.

And last: if you really want to stay in science you must be aware that you are not able to plan your career as you possibly are in industry. So you need enough enthusiasm and be able to tolerate a high level of frustration and most important you must be able to meet challenges within a short time like Dr H already pointed out - so if you really think this is not the right place to do your PhD you should start to look around, and if you think you can manage the tasks and survive the lab you should start to work hard to be the one of three PhD students who make it ;-)

and now, an Engelbert Humperdinck's 60's classic:



(a kind of twisted love song from a fruitfly to dr H):


Please release me let me go
Cos I don't want this anymore
To waste my life would be a sin
Release me and let me fly again.....

...

you shld give me a plus for this, dr H.....

What do you call a 'chit-chat' lizard in English? That's the Indonesian slang name for a common little lizard.
iPhone

I'm sorry, I think this is just a myth.  Do the math, instead of repeating a rumor.  Your PCR reaction has a final concentration of magnesium at about 2 mM.  If you have your primers at 10 uM in TE, and your final primer concentration is at 0.5 uM, then you are doing a 20:1 dilution. TE has 1 mM EDTA, so the final EDTA concentration is 0.1 mM (0.05 mM from the addition of each of two primers).  So, the final magnesium concentration ends up at 1.9 mM, rather than 2 mM.  This is inconsequential.

To agarose manufacturer,
we are currently trying to increase awarenesss of the fact that product advertisments are really annoying in the Methods subforum. The subforum is a molecular biology grade, and you should post info about your free trial to Free Stuff or other Products and Vendors subforums where it belongs. This allows the clear distinction from methods-related questions and good reputation of your company. The computer I'm writing this on is powered with nuclear energy (probably).
I would love for frequent spammers (not you particulary as you are yet not frequent) to try posting to the right subforums. If you would like to offer us something, please don't post it here (and DON'T message us) and we may try it. We have a limited patience, so it is based on first come, first "vote this post down" basis. We are also often outside the United States, so limited shippings makes us sad.

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