Tuesday, November 22, 2005

DNA restriction enzyme analysis workshop

Looking at the title, you might have wondered whether am I holding it or sth. lol. no.. In fact, I went to it last Friday, 18th nov. It was held in the Singapore Science Centre. I thk I can roughly remember the details of it:

Introduction: This workshop is mainly about restriction enzyme analysis and gel electrophoresis. In restriction enzyme analysis, specific enzymes are used to digest DNA, thus resulting in banding pattterns for comparisons and analysis. Gel electrophoresis separates the digested DNA fragments in an electric field to produce bands of DNA ( you can try guessing: DNA moves to the +ve or -ve side of the field?) . This technique is also the basis of DNA fingerprinting - often used in screening for diseases and solving criminal and paternity cases

09:30 - 11:30 a.m.: The workshop starts. Of course, the trainer introduces herself and went to introduce some facts abt DNA and also the micropipette and other apparatus in the lab. One micropipette costs abt S$500 and a bottle of agarose powder costs I thk S$800 if I'm not wrong. Ok, so then, we sort of practise with all the apparatus especially the micropipette. We did a run on sample dyes to get bands and we do case studies. After all that, we started on the real thing coz time will be needed for incubation in 37 degrees celsius environment in which that time will be our break. Okay, so it was like a case study thing again in which lots of bottles restriction enzymes were broken but onli 4 bottles were unbroken and we have to analyse and identify each restriction enzyme in the bottles. The 4 different restriction enzymes used by the company in the case study are EcoRI, HindIII, BamHI and NcoI. Lambda DNA (it's a bacteriophage which is a bacterial virus) was used as it is only 48, 502 bair-pair long, so is good for analysis. So four microtubes were labelled 1-4 and inside were different restriction enzymes. But of course, we have to make the agarose gel first. After the agarose was done, we have to wait for it to set. So in the mean time, we prepared the digestion mixture which is a mixture of the restriction enzymes, dye and DNA. The tubes were centrifuged to ensue that all the liquid in the tube settles to the bottom of the tube. And the tubes were placed in a 37 degrees celcius incubator/ water bath (I thk it's incubator) and time was also given to the enzymes for them to digest. Then, time for break.

11:30 - 13:30 p.m.: Break-time. Me and my friends had lunch at Macdonalds' and explored the science center. It was realli cool. I mean, its been like 4/5 years since I've been there haha. There was this human body exhibition, some physical science exhibition and of course we explored a little of the DNA trail since we had to go back for the workshop.

13:30 - 15:30 p.m.: The workshop continues. Now, the samples are ready for gel electrophoresis. 10 micro litres of each samples were loaded into four different wells in the agarose gel. We had to let it run for 45 min. In the mean time, we have to interpret restriction maps and calculate the sizes of the different fragments that each restriction enzyme will cut at specific sites of the lambda DNA. Then we did a drawing on the positions of the DNA fragments on a diagram of an agarose gel as these will be seens as bands on the actual agarose gel after it has been appropriately stained with ethidium bromide (highly mutagenic substance, must be handled very carefully and appropriately) and viewed. We watched some videos and very soon, the 45 mins were up. The gels were then taken for staining by the instructors and in the mean time, we did some more case studies. Soon after, the results of each group were printed and the enzymes were identified. The first tube is just the DNA sample with no digestion coz there was actually no enzyme inside. The second tube contains EcoRI, the third - HindIII and the last one is a mixture of EcoRI and HindIII. Yep. Then we did a group survey and it was the end of the workshop.

15:30-16:30 p.m.: One hour of exploration once again before it was time to leave for school.

It was a really fun and enjoyable experience althogether especially with the handling of the micropipette and other apparatus although I've handled a micropipette before when I was in secondary school, this time, I had longer time with it and more usage too. Not only that, I gain more knowledge about DNA profiling from the case studies. The case studies were realli interesting too. The videos shown were also very educational and enriching for us. This workshop reinforces my skills and knowledge in this area and not one minute was wasted ;). It was realli cool to be in the lab and wear the lab coat as well, makes all of us feel like scientists, haha.

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