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Doug and Brian with one of the shark pups

So, as you can probably tell from my Hawaiian shirt and crazed smile, today was the last day of classes for me. How did I decide to celebrate? Remember last week when I posted this Tweet

Well, the backstory is that a six-gilled shark washed up on a beach in North Saanich (just north of Victoria) and Dr. Julia Baum and Dr. Francis Juanes from UVic went with some of their labs to perform a necropsy. Turns out the shark, which was about 4 metres long, was pregnant. After seeing my Tweet, Brian Timmer from the Juanes' lab contacted me to let me know THEY HAD SOME OF THE BABIES AND I COULD COME AND SEE ONE! If you want to read about the necropsy, go here. There are some pretty awesome photos.

Fast forward to today, the last day of class, and I got a chance to see one of the pups. It was awesome!

Well, the dog days of summer are over and school is about to begin again. This year I thought I would address some concerns and difficulties that students have had in the past. You can use this as a handy reference guide for all of my classes.

1) Go to class and pay attention! This is a big one. Most of my exam questions come directly from lecture material, so it is critical that you go. If you have taken classes with me before, you will be familiar with my posted notes. What do you notice about them...there is not a lot of information. They are designed to lead you through the lecture, but not be a comprehensive overview of what is going on in class. If you are having difficulties taking notes, check out the UVic Study Solutions webpage.

2) Read through the notes and assigned readings before lecture. This will help you to follow along.

3) Do all of the practice problems. For all of my courses, I post old exam questions. Sit down and do these as if they are an exam. Just flipping between questions and answers will not help. If the course has online or textbook questions assigned, I highly recommend doing these. A very small number of students currently complete these problems, and yet on CES (course evaluation surveys) students always want more practice questions. Here they are, come and do them!

4) Consider forming a study group. This is especially helpful for courses with multiple choice exams. You can all contribute questions to test each other. The bonus here is that when you are making questions, you are approaching the material in exactly the same way I am when I make up your exams. You may get incredible insight into the inner workings of my brain this way.

5) Utilize office hours, feedback links on CourseSpaces or email if you are falling behind or need clarification for something covered in lecture. I try very hard to be helpful, but you will need to work with me. If you tell me " I don't get any of this", I will not be able to help you. Work through it as much as you can on your own and then come to me with specific questions or problems. We can use this as a starting point and avoid needless frustration on both of our parts. I will be able to be a lot LOT more helpful if you do a bit of legwork prior to emailing me or coming to see me.

6) Speaking of email, when you are emailing me a question, make sure you put the course number and a brief description in the subject line. Also, don't start it out as "hey dude". While I'm not a particularly formal person, many of my colleagues are, as are most employers, so get into the habit of good email etiquette. Dr. Cheryl Hill posted this handy guide on her Twitter feed (@HillPhD).

 

 

7) Try to get at least 12 hours of sleep a night and drink plenty of water. Let me know if this helps. I have never actually managed to achieve it.

We set up some grids of our protein for both negative staining and cryo-EM. There are a different types of grids, but a typical example can be seen here with a pen for scale. We were using copper grids coated with carbon for the negative staining, and gold grids with gold foil (UltrAuFoil) developed by Dr. Lori Passmore and Dr. Chris Russo here at the LMB for the cryo-EM. The grids have to be made hydrophilic, which is done by either glow-discharge, shown here, or plasma treatment (see my recent Twitter post for this). Either way you get a cool looking violet light. Very scientific. Samples are added to the grid and this is basically where the paths diverge for the two methods. For negative staining, the sample is stained with uranyl acetate. This allows you to see the shell of the protein, but not internal structures. All of this is performed at room temperature, as is the actually microscopy. For cryo-EM, there is no stain, and internal structures are visible. As the name implies, very
low temperatures must be maintained throughout the cryo- process. Samples are added to the grid and blotted so that they are in a very thin film of water. The samples is then plunged into liquid ethane (yes, liquid ethane, smoking is strongly discouraged here) at -181C. You can see the special container for the ethane here, with the small well in the middle holding the ethane, and the surrounding bath containing liquid nitrogen. At the top you can see the robot in action. Protein is spotted, blotted and plunged into liquid ethane. I have also stuck in a picture of me checking out our negative grids (which was not, unfortunately, as massive a success as I was hoping for).

This has been a very busy week, which is why I haven't had a chance to update before today. I have seen great talks on structural biology from Patrick Cramer (Max Planck), Elizabeth Villa (UCSD) and John Schwabe (University of Leicester). I will try and put some of this material into BIOL 225 and BIOC 300A. Today there is a talk from Richard Henderson, who won the Nobel for his work in 2017 that I am pretty excited about. To top it all off, Right Said Fred is playing for free tonight in one of the local parks.

We have been doing a lot of classic biochemistry experiments this week, as well as

some crystallography and baculovirus work. I will talk about that baculo work in a later post when I have more time. Crystallography is not being used as widely at the LMB as it probably was in the past, as they are trying to maximise use of the cryo-EM facitilities. The facilities for crystallography are not that different than they were in Frank Sicheri's lab in the 2000s. There is more automation, but there is still a lot of manual screening, so you still have a great opportunity for punishing eye strain. I don't know if it will load properly, because I have been having some technical difficulties, but the video here shows the robot revolution in progress! Quiver in fear feeble humans!

Also, there was a football game that pretty much everyone, including myself, was watching.

Friday was our cryo-EM day. The set up itself is pretty complex, and it took a while to get things up and running. The challenge revolves around getting everything in alignment, since the accuracy of the machine is so high. By the end of the day we were on our way with the collection running until Monday. You can see the tired crew in the photo. Proteins are distributed in a gold grid that has, in this case, a very thin gold foil with holes in it. There are several hundred squares on the grid, and several hundred holes in each grid square. We set up to count more than a thousand holes. A sample image is displayed on this page. The dark clusters are protein complexes. You can see that there is very little information in each picture. In the next step, the data will be compiled from all of the images and processed. Hopefully, at the end, a beautiful high resolution image will emerge!

The Passmore lab's focus is structural biology. They utilise a large number of techniques, including cryo-EM, which is an excellent tool for analysing the structure of large complexes. This video features Drs. Passmore, Scheres and Henderson (2017 Nobel Prize in Chemistry winner) from the LMB explaining the technique. There are several projects running in the lab. These projects all revolve around multi subunit complexes. The main focus is on proteins that bind to the poly-A tail of eukaryotic mRNA and a complex involved in DNA repair that is linked to Fancomi anaemia. Yesterday was a big purification day. It was great to get back on the bench after a multi-year hiatus. It is all slowly coming back to me. I even managed to load a gel without injuring myself or any others. The UV trace above shows that the purification worked well.

On a more personal note, my badge has gone from temporary to just a normal pass. On the down side, I look like some sort of deranged criminal in the photo.

Also, this computer is set to proper UK English, so all of my "z's" are being changed to "s" in words like analyse and utilise.

So, the adventure starts with a walk along the Cambridge Busway to the MRC-LMB. The idea behind the busway is that there are two dedicated bus lanes, separated by a 15 cm or so curb from a cycling/walking path.  The trick is that the buses move at about 100 km/hr. It is a bit unnerving the first time one passes you. This video gives you an idea about how it works.

The LMB is in an amazing new building with a huge atrium and labs running along the sides. From a history point of view, there are loads of interesting artifacts from some of the famous labs and experiments spread throughout. Here is a timeline of some of the important discoveries from the LMB and here is a catalog of some of the historical scientific models on display. It is hard to imagine modelling a structure without computer software, but the amount of detail is remarkable. The photo here is one of Kendrew's models of myoglobin, the first protein structure solved.

I have also been upgraded from "Visitor" to "Temporary" on my first day. This seems like significant progress! I will tell you about lab life in a subsequent post.

As you probably know, I have won several important awards for not updating my blog, but this time things will be different. I will be working for a month in Dr. Lori Passmore's lab at the LMB in Cambridge. It has been some time since I was on the bench, but I'm sure it is just like riding a bike. Actually, that is something else I haven't done in many years but plan to take up while in town.

This spring, the UVic Department of Biochemistry and Microbiology hosted Dr. David Lewis, Director of the Western Sydney Sexual Health Centre, as a Lansdowne lecturer. He gave two great talks on antimicrobial resistance in Neisseria gonorrhoeae, the bacterium that causes gonorrhoeae. The timing was very good as the World Health Organization had just listed antibiotic-resistant N. gonorrhoeae as one of twelve priority antibiotic-resistant bacteria. The topic was making a big splash again yesterday  (Nature, the Guardian, BBC) as a new paper on the topic by Wi et al. is in the current edition of PLoS Medicine.

You should check out this fascinating article by Browne et al. (2017) about the transmission of commensal microbes. We talk a lot about how pathogens get around, but what about commensals?

There are a somewhat limited number of routes, including fecal-oral and mother-child, either during childbirth or breastfeeding, but how do the microbes get into the breast milk? We hear a lot about probiotics, but how do they work and do they effectively establish long term colonization?

Finally, what are the effects of perturbations such as antibiotic treatment? There is evidence this may lead to extinction of some species within a population. On the flip side, is there a future where we use biotherapeutics, introducing bacteria to treat disease, rather than wiping bacteria out?