Christos Katsifis, Week 1

           I have luckily dodged the stereotypically rainy and dreary English weather; it has been slightly north of 80 degrees and the sky has been cloudless. The only English stereotypes to truly be upheld here in London are the frequency of cursing, an obsession with soccer, and a concerning prevalence of alcohol, but besides that, I have been very surprised. The food is not just fish and chips, the population is very diverse, and the people have been very friendly. (Side story: There is a large, beautiful park located about a mile away from my apartment called Hyde Park. There are soccer fields, rugby fields, a massive royal shrine, but there is also a wide, cobblestone path bordered by tall trees and small park benches. I had spent a majority of my Sunday before my true lab experience reading on one of those benches. About an hour in to my stay, I was approached by a homeless man asking, "Ey mate, would ya mind offerin me a fag?" My immersion within my book was disrupted, and I was also plainly really confused, "Excuse me sir?" The man repeats the question in different context, "You know, a cigarette mate." Obviously, I apologized to the man for I did not have any, but he sat down on the bench regardless. He continued to distract me from my reading for the next half-hour or so, talking about life in different contexts -- American and English -- and debating over the most prolific striker in the premiere league. He was very friendly!). 

         Perhaps my favorite part about London is that everyone cuffs their pants like I do; I must look more European that I anticipated, as everyone here dresses like either me or Omer (Turkey is very European, while also arguably being in Asia).




There is also something incredible about the grass here in England. It sounds rather silly, but the grass is startlingly perfect.

     





          The first two days I had arrived in London, my father and I explored a couple of the remarkable museums only a block away from the apartment. We had visited the Museum of Natural History on the first day. In the Museum of Natural History, there was a human evolution exhibit. My father has told me for as long as I can remember that I look like a Neanderthal. So given the opportunity within the exhibit, I took a picture for comparison. 

I personally do not think I do, but it is, of course, not up to my discretion.

            My first day at the lab went pretty well. I had been instructed to go to the Sir Alex Fleming Research Building at the time of my choice and ask reception for the name of my PI, Dr. Tony Nolan. I was slightly disoriented by the "time of my choice", so I decided on eight o'clock. Unfortunately, I was not aware that he, nor anyone else in my lab arrive until ten thirty; the receptionist and I had a couple of laughs about the situation. After grabbing a coffee from the local "Paul's"-- a fantastic french coffee shop serving homemade pastries and sandwiches for the price of a McDonald's Big Mac-- I sat in the lobby/lunch area of the SAF Research Building. I had a book to pass the time.
         
         Dr. Tony Nolan stands at around 6'3", and has a stern complexion that not even Mr. Onion could match. After a brief introduction and inquisition of my impression of London, I was informed of my four-page, single spaced report on the function, capability, and ethics of gene drive systems that he expected upon my arrival the next day. The rest of my day was occupied by lab safety inductions, insectory inductions, and independent research on two strands of mosquitoes (Anopheles gambiae and Anopheles funestus- two major malaria vectors).

This is a picture of my foot and I, watching the sun rise while finishing the gene drive report.

         During the first week, I was assigned multiple tasks: the complete process of rearing mosquitoes, micro-injection of transgenic DNA in to mosquito embryos, and the optimization of water culture for Anopheles funestus (the more difficult of the two species). 

        I worked with Mariana to learn the process of rearing mosquitoes. She is a Brazilian technician who helped me bleach mosquito eggs, hatch them in to trays, sex and sort the pupae, and then organize the transgenic mosquitoes from the wild type in to individual cages by fluorescent screening. 

       I worked with Dr. Andrew Hammond and PhD student Kyros Kyrou to align and micro-inject mosquito embryos. Upon being assigned to work with Dr. Hammond for the day by Dr. Nolan, Dr. Hammond plainly told me to "tell me what you know about gene drive". I told him what I thought of gene drive systems and regurgitated the function and use from the four page paper I wrote. (Gene Drive systems are an integrated piece of transgenic DNA, containing a sequence coding for a CRISPR/Cas9 complex. Once the transgenic DNA is fused in to the mosquito embryo DNA, the Cas9 endonuclease cleaves the target sequence -- usually the homologous sequence in which the CRISPR/Cas9 coding divides -- and prompts homologous DNA repair, which then copies the CRISPR/Cas9 sequence in to the opposite chromosome, thus causing a homozygote. This helps force the dispersal of a specific trait amongst a population.). Dr. Hammond and I continued to converse, more so a lecture, upon the capabilities of gene drive and its current applications.The alignment of mosquito embryos was relatively difficult. As the embryos are borderline microscopic, the alignment must be done under a dissecting microscope. I used a paint brush to organize the bean shaped eggs, with the seemingly fatter end facing forward, along a piece of paper with a sponge on top to prevent the dehydration of the eggs. The fatter end is the developmental head of the mosquito, while the skinnier part is the abdomen. Since the transgenic DNA interferes with a gene involving female fertility, the DNA was injected in to the abdomen end. The micro-injection was done in an electron microscope. The needle was barely visible, as are the eggs, and with fine, hydraulic tuners, the needle is guided to barely penetrate the barrier of the embryo. I only sacrificed a couple of embryos.

        I was assigned one preliminary project, which is the optimization of the water culture with the mosquitoes. With a bit of research, I determined the optimal conditions, as the mosquitoes proliferate best during harvesting season for rice. Rice secrets Vitamin B, Vitamin E, phosphorus, and magnesium in to the surrounding water, so with a test culture of the rice water solution, the mosquitoes developed very quickly, compared to that of the normally used mineral water (21 days from egg hatch to L4 larvae/pupae versus 4 days). I have yet to learn of my next project. 

      I am hoping to work with Dr. Hammond in the creation of sgRNA's for CRISPR/Cas9 complexes, and possibly experiment with CRISPR/Cpf1, which allows for multiple sgRNA's, in the upcoming weeks.
 
     During this time, I am incredibly appreciative for my roommate, Zoe, for she came back from her weekend trip from Frankfurt just in time to let me in to the apartment because of forgotten keys (the first, and last time it will happen). I had eagerly rushed out of the apartment earlier on that Monday afternoon to play soccer with my lab mates, Dr. Nolan, and some British friends of theirs. The British players were truly hilarious, with loud grunts of profanity resultant of any misfortune. Dr. Nolan was pretty good; I tried to stay on the opposite side of the pitch to avoid somehow roughly knocking in to him. I do not think an aggressive infraction would serve me well. 

    Thank you Mr. Sham and Dr. Peretz for your help. I am looking forward to seeing you Mr. Sham on the ninth.