24 November 2010

My First Scientific Report

Hello blog! It's been quite a while since I last scribbled on your dear face. I am to blame for it really, it's not you...it's just that I'm spending my time 'hanging out' more with my lecture notes since I came back. It's been a quite gruelling month to be honest, with lectures coming thick and fast, I need all the time I have to make my notes and prepare for the next one. But Alhamdulillah, its been okay so far. In fact, I'll tell you and the others about my first experience of preparing a scientific report. You know...the ones REAL scientists do for a living. Ready? here it is...

Genomic Library Production of Bacteriophage Lambda via Cloning and Identification of Gene Fragment

So, that's the title. Then comes the Abstract. What's that you say? Oh, its basically a summary of the whole experiment. In a paper, it comes right in the beginning after the title but in terms of preparation, only once the whole experiment is complete can you proceed to make it. Kind of like a synopsis to a novel, like the ones you're used to making in the days of reading KOMSAS texts and stories like the Prisoner of Zenda....ah PMR, good times..good times. Anyway, as in any summary a good understanding of the whole experiment and its outcomes is needlessly essential in making a well written one. The text also had to be short enough so that anyone reading it will already understand the whole process and determine whether it is suitable or not for his/ her specific research. You see, scientists rarely do have a lot of free time so they'll always try to scheme through where possible...not working for me yet but I'll get used to it someday.

Then comes the Introduction. Well, basically its an insight regarding the particular biological aspect were going to be talking about. In my case, explanations regarding the bacteriophage as to what it is, why was it used, the genes within it, the cleavage points within, the restriction enzymes used...anything concerning the topic can be included really, but as always care has to be included in making any particular statement. For example, if you say that "bacteriophage lambda is a molecule consisting of 48kbps and 12 single stranded nucleotide termini in its linear state", back it up. Quote at the end of the sentence as to where did you come up with this piece of information just to make your paragraphs fancier. If you did, then well done and you get marks for it. If not, it'll be treated as a fallacy and could possibly be rejected in the scientific community, a total waste when you consider how man hours one usually spends on these type of articles. In the real world, there will consistently be scientific forums to review newly published research articles and they are a lot more unforgiving than what some lecturers can be. Even the source has to credible enough. In most cases, data is obtained from scientific journals and texts. Lecture notes are certainly a no-no so please under any circumstance do not put them in quoting your lecturer as the source, no matter how temptingly easy this may seem,hehe :)

Moving on to the Methods. Basically, its very easy to write as much as you want in explaining the steps you took. But here comes the hard part. Publishing a scientific report is expensive as each page can cost in excess of 300k of pounds, so its vital that the paper focuses more on the outcomes than the steps taken to complete the experiment. However, bear in mind that the methods must also be sufficient enough to allow anyone to repeat the whole experiment successfully, which in certain cases they do in order to verify the outcomes of certain experiments. So its quite a tricky business of balancing what to put in and what's not, but as always practice makes perfect. Oh, and its also important to explain the reasoning behind each particular step as it makes the results more accurate and easily said, believable. Methods are usually separated to sections with reference to the aim of doing a particular method instead of a timeline. For example, section headers may be titled as "Restriction enzyme digestion","Gel Electrophoresis","Plating Bacteria" instead of "Week 1","Week 2"...got it? so yeah, it makes it a little bit more difficult since the steps are not exactly done all at once but it does make it clearer to the reader. Again, emphasis on making it easy for the reader... :)

Now comes the cool part, Results and Discussion. In this section, one is expected to show the results of the experiment in the form of figures complete with their legends (no, not the one in the classical history segment, mind you!). Legends are basically short descriptions as to what the figures refer to. Simple enough really especially when you have space to explain each and every step of the way. So then, here's my sample to share with you dear readers.

Once upon a time....a bacteriophage lambda was cut into fragments using the restriction enzyme Pst1. To confirm this has happened successfully, samples of the digested molecule were subjected to gel electrophoresis with the presence of controls being the undigested samples.

(apologies for the small picture, increasing its size will only cause the resolution to decrease where by then it defeats the purpose of putting it in in the first place) As you can see in well 3, the genome was cleaved into 27 different fragments while in the control in well 1 is still in one piece, proving that digestion was successful. At the same time, the gel was also cleaved using restriction enzymes EcoR1 and Mlu1 with HindIII digested lambda acting as the DNA marker to determine the size of the fragment, as seen below.

These are particularly essential in determining the position of the specific cloned fragment via Southern Blotting, but more on that later.

Next, we then insert one particular fragment from the 27 available into a plasmid vector. This is later transformed into an E.coli cell to allow the cell to grow and hence, clone the specific fragment along with it. The next challenge lies in the determining which cells have took up the recombinant (altered) plasmid and which ones still have the unchanged ones. This is done via blue white assay in an ampicillin plate. Growth on the plates means that the cells have successfully taken up plasmids while in the blue white assay, blue colonies proves that the lac z' gene within the plasmid is still intact, hence proving that the plasmid incorporated was not the altered one. White colonies on the other hand proves that plasmids containing the gene fragment was successfully cloned. Sorry if this is a little bit too confusing...

Upon completion, samples from the white colony was again digested using Pst1 to release the cloned fragment. This is later run on an agarose gel with standard DNA marker to identify which fragment was cloned out of the original 27 and what is the size, again as shown below.

My fragment is the one at the end and as we can see, the size of it is roughly around 5kbs. Using certain appendixes provided, the precise value of this is determined to be 5.007kbps. However, from this, it is still not impossible to determine the precise position of the fragment within the genome and here is where the Southern blot becomes vital, as shown below.

The blot shows the labeled fragments of EcoR1 and Mlu1 digests. This proves that the position of the lambda fragment cloned is within these two fragments position and hence overlapping of both with the help of appendixes shows that the cloned fragment is from the part of the bacteriophage which codes for integration, excision and recombination of the genome.

Once all that is done, references must be quoted in the end and any acknowledgements made are also welcomed. Again, this must follow a given format, specifying who are the authors, the publication year, what page and many more things to consider. Quite a hassle really, but like I said, the references mean everything in validating the authenticity of the experiment so just live with it, keep calm and carry on... :)

All in all, it took me roughly a month to complete the experiment and prepare the report. Trust me, its not an easy task to complete and to be honest, I only fully understood the whole experiment 3 days before the deadline but thankfully I nevertheless managed to submit it on time. This experiment counts for quite a significant portion of my final marks for the year so I'm hoping for the best. It's certainly been one cool experience and I anticipate many more in my years to come as a scientist. Sorry for any scientific terms being less understandable but trust me, with a little bit of effort you'll understand it as well as I do in no time. Cheers!

P.S: to those taking their SPM exams (e.g: Yoyong and Yin) , all the best!


Musafir Melayu said...

baku, what course are taking actually?

oh, it's been a while since your last post.

thanks for sharing a bit of your life! :)

Musafir Melayu said...

*are you taking... --"

Farhan Baku said...

I'm doing Biotechnology, You know...play with living stuffs like cells and protiens, altering them for your own interest,hehe :)

Nabila Azureen Azis said...

salam. thanks for the wish. wow, how scientist you are. :)

Farhan Baku said...

marks just in...Alhamdulillah, I got 86.5 percent! not too bad really for a first go,hehe :)

Nabila Azureen Azis said...

waaa. for this, may i convey the first class of congratulation to you. hihi, congrats, that's a really high score :D