11/20/13

This week, I got to officially start on my very own experiment!!!

 As mentioned in my previous post, my experiment revolves around potassium. My mentor and I decided to incorporate this very metal via the agar plates. Rather than the usual set of ingredients used to create the standard agar plate, I added a particular amount of potassium bicarbonate to the mix in addition to the regular components. As shown in the picture to the left, I prepared 3 separate mixtures--one of which is the control medium.

(Here is a bonus picture! :D) This is me in the compete lab gear head to toe. (except my sterilized clogs aren't shown here). In this image, I am preparing plates (moving the spore onto the agar plates) as mentioned in the post below.

11/13/13

        Due to the fact that now I have basically went through the general process of growing the mushroom product they produce at Ecovative, my mentor told me that we would be moving on to a new project. This time, she gave me full rein. My next assignment/opportunity was to create my own experiment. She asked me what I would like to experiment on/about and that once I had a set topic, I would get to carry out the experiment, like how an actual scientist would him/herself. First, I started with a word bank of vague and broad ideas that I was interested in and that which would be feasible for me to carry through. After reading several papers that she printed out for me and researching on the web, I decided to base my very first project on the workings/effect of potassium on the growth of mushrooms and also on the inhibition of molds/bacteria of the growing strains. This topic, my mentor and I agreed, would give me various pathways to take even after performing the experiment. For instance, not only would I be able to do the actual experiment, but also I would be able to get to use the Dinoxcope program (program that allows one to use the microscope via the computer screen) to analyze and save my results later-on.

      Aside from deciding my project, just to practice and better my plating skills, my mentor left me all by-myself to 1) wipe down the hood, 2) get the materials 3) perform plating plates 4) clean the hood. In short, she basically left me to do my own things while she took care of her other projects. I felt so accomplished, content, and most importantly: independent by the end of my internship. I can't wait for the coming weeks when I get to actually start my project and see how it takes shape!

11/06/13

In addition to the things that I did last week, my mentor and I also set up agar plates around 4 different places in the lab to see what kinds of molds are in the atmosphere. I placed one agar plate near the sink, one near the cookers (the pressure cookers), one near the incubation rack, and one near the compressor shelf. Coming in today, I was really curious as to see the different kinds of molds that should have formed over the past few days while I was gone. To my great dismay, not that many molds or bacteria formed on the plates--this is a good thing in the company's stance as that means their lab for the most part is a sterile environment--which is what they aim to attain. However, the agar plate next to the pressure cookers showed three groupings of mold that had begun to take form. In addition, the agar plate that was placed near the sink exhibited a blob of bacteria. My mentor taught me how to tell the difference between molds and bacteria. Contaminants that look watery and slimy are most likely to be the latter group (bateria) while dry and fuzzy substances the former (molds). After seeing these, I finally understood the reason why my mentor stressed ever-so-much the concept of sterilizing every single tool and person before conducting experiments. Even in this seemingly-pristine environment, there are still contaminants in the air that can harm the products.

This is bacteria

These are 3 molds

 As well as that, today I was able to take home my first test-product-mushroom-packaging product!! After I took the picture of the teddy bears posted in my previous blog post, my mentor and I put them in the dryer to absorb all the moisture and thereby prevent the mycelia from further growing and fruiting mushrooms. Had these objects simply been left out in the humid atmosphere, they would turn into something like the picture below.

<-- The protruding object circled in yellow is the beginning of a mushroom. Left over time, this would eventually grow into the mushroom we know and imagine today.

I got to take these three products back with me!!

 Upon showing my first test products to my friends, they all believed me when I lied and told them it was styrofoam. When I told them it was actually made of compost materials and mushroom mycelium, they were very surprised. :) 

10/30/13: MUSHROOM TEDDY BEARS :)

Last Wednesday, I continued with the next step in the process of making my very first mushroom test-product at Evocative. In the previous post, I shared my experience in putting together a bag of growing medium (compost) for the mushrooms to grow on. After these bags show signs of inoculation (also as stated in the prior post), one would open them up and throughly mix them together--and this is exactly what I did. 

After my mentor and I decided that the two bags we had left to inoculate were sufficiently ready, she informed me that my next procedure would be to pour the compost, now covered with newly-grown fungi, onto a flat surface and to completely mix it so that it would give off a homogenous appearance rather than only the surface portion of the bag containing the white mycelia. (this is shown in the picture to the left)

After accomplishing this task, I got to choose the type of mold to pack this compost in--I went with the teddy-bear shaped mold. As mentioned many times before, the mycelia (white things that appear during inoculation) acts as a glue for the compost materials. Thus, when the mycelia-infused medium is tightly inserted into a plastic mold, over a time period of three to five days the agricultural waste transforms into a durable material. Depending on the type of strain of mycelium used in the prior step (the step that I explained in my previous post), Ecovative is able to make a variety of different kinds of the final product. For example, one type of strain would produce a water absorbent product while another would produce a dielectric final packaging product. In this way, the company is able to meet the requests of many different types of companies and each of their products (ie: computers, wines, etc).

Before

After 3~5 days

10/23/13

    Other than check up on my agar plates last week, I also had the chance to put together a bag of growing medium (compost) for the mushrooms to grow on. This is the very first step Ecovative scientists perform when creating their mushroom product. What one does to create this medium is first insert into a sterilized bag a sufficient amount of straw, corn cobs, etc... and on top of that add pure grain spawn on top. Next, one throughly mixes the two ingredients together to fully inoculate the grain. Then the bag is tightly sealed and left to grow.

                  Unlike flowering plants, all mushrooms grow from tiny spores rather than seeds. (Plants that grow from spores are called fungi). Due to the fact that mushrooms have no chlorophyll to obtain nutrients by they must get their nutrients from organic matter in their environment--this is most often called compost, a combination of various materials like corn cobs and straw. This goes back to the point I made in my very first post concerning the eco-friendly production at Ecovative. This company is planet-friendly even in the sense that the ingredients needed to create their products are what many consider "waste." Ecovative uses the surplus of leftover corn cobs, straw, seed hulls, etc... that are left over after harvest season to create their compost for the mushrooms to grow on. A full-grown mushroom will produce many many spores (ie: 16 billion) and they must then be collected in a sterile environment, like the lab Ecovative scientists work in.

 Thus, coming in this past Wednesday, I found the two bags of compost I put together fully cultured. When before the bags looked like a batch of straw compressed together, now it gave off a white appearance, indicating that the fungi had grown as predicted. As shown in the picture to the left, the white mycelia colonized the entire compost--this visible growth of the fungi can be used as an indicator of its life cycle stage and health.

Each week, I advance through the steps Ecovative scientists perform routinely to create their various products and each week I can't wait to see what the next procedure is! I am also really excited to see how my very own Styrofoam-like mushroom packaging product will turn out!

10/16/13: Another day at the Mushroom Factory

         Last Wednesday I finally got to see the results of the agar plate onto which I had transferred the Mycelial Culture. In my previous post, I addressed some concerns in which I feared I may have contaminated a majority of the very plates. However, to both my mentor and my delight, we found that most grew without a mold in sight--only about one or two plates showed traces of bacteria.

   Expanding on last weeks project, the agar media that was used to grow the mycelium is not a fertile ground for growing mushrooms--the fruiting bodies of the fungi that produce spores. However, the mycelium culture grows quite nicely on it and thus the mycelium that grows in petri dishes (like the ones below) can be used only to inoculate a substrate where it can further develop.

As you can see in this picture (to the left), the white mycelium, resulting from the spore placed last week on the fresh agar plate, is now visible. This is perfectly normal and tells us that everything went according to plan. The first signs of mycelial growth gives off a white fuzzy looking appearance --> called rhizomorphic growth. It is a sign that the mycelium will most likely fruit well. Most of the plates were completely colonized as shown by its completely white semblance. As mentioned before, any other abnormal substance, ie: mold, is a clear sign of contamination. (A contaminated plate will never produce mushrooms)

This picture to the left is just the underside view of one plate held to the light. You can see the spore placed that I placed the week before with a sterilized scalpel in the center. The culture grows in a ring-like shape and this is clearly shown in the very picture. Had I been here more often than just once a week, the normal procedure would be to note the growth of the mycelium by measuring the diameter of the ring formed.

Each week here simply gets more exciting by the week and I can not wait to see what else is in store for me to put my hands on throughout the year!

10/09/13: DAY 1

Last Wednesday, as soon as I arrived to Evocative, my mentor put me straight to work. We headed immediately to the lab to make agar plates that the mushrooms grow on. Before entering, I had to completely sterilize myself. To do this, I had to exchange my ugg boots for sanitized crocs, wear a lab coat, put on top of my lab coat clear sleeve covering for each arm, wear gloves, put on a hair net, and to top it all off, I had to wear a face mask. All this to make sure no bacterium contaminates the products. Even before I entered the lab I was already in awe.

The big thing that I learned on last Wednesday was about the very topic of contamination and how to avoid it. Even when working in the laminar flow hood transferring mycelial culture on the cooled agar plates my mentor constantly called me out for waving my arm—which was protected by a lab coat and a protective sleeve over it and misted with an alcoholic solution—over the plate that contained the mycelial culture. She told me that each time I would either have to open and then close right away the petri dish when transferring the culture to a fresh plate or find my own way that does not involve waving back and forth my arm over the colonized plate. Courtney informed that each time she makes plates, she makes at least 20 so that she knows the results she obtains are correct and not because of some particular mutation. On that day, I made about 24 plates and for more than half I was called out for. Tomorrow, I will get to see the results of the culture I transferred and I expect very few to have turned out normally. Most I predict will have soiled and formed a mold-like substance on them. By the end of my stay there, I found myself cut off as my ride had arrived. There were simply an infinite number of things for me to do that my mentor had instructed me to simply cut her off when it is time for me to leave. I can not wait to get back to Ecovative and immerse myself amongst the busy scientists, engineers, designers, and chemists that are too busy to even realize that they’ve missed lunch tomorrow!

10/03/13: My First Mushroom Visit

Last Wednesday, I had my first visit to Ecovative and to my great delight found that it was everything I hoped to find and more.

Upon first entering the doors to the company, I was met with a contract that I was asked to sign, regarding confidentiality of their mushroom productions. They explained to me that it was just in case I don't decide to start a mushroom packaging company of my own that would challenge theirs. :)

My mentor, Courtney Hart a research scientist, showed me around and what I saw behind the double doors took me by surprise. What had just moments ago been an office place filled with computers and desks had transformed into a very spacious and factory-like place. Everywhere I saw there were enormous machines that played a role in the whole mushroom material-making process. One-by-one Courtney explained to me what they were used for and I remember just standing there in awe of everything around me. It was my first time getting to observe the behind-the-scenes workings of a "science company" and what I saw left me open-mouthed and with a broader view about the the applications of the very subject of science in the real world. When last year I was limited to the lab in both my biology class and my internship, here I given full access to much much more.

After the preliminary tour around the factory/company, my mentor showed me the place where I would be spending most of my time: the lab. Here, I found that before entering I would have to change my shoes into sterilized crocs and wear various gears (ie: lab coat, hairnet, etc) to make sure nothing contaminated is brought inside the lab. Courtney explained to me that in this very lab, they grow their home compostable material. Giving me a brief rundown of how the company basically works, she mentioned that Evocative first buys massive tons of leftover agricultural byproducts (ie: corn stalks) and combines them with root structures from mushrooms called mycelium--this which is used as a natural self-assembling glue. The material is then allowed to grow in the dark and left to self assemble into a solid mass. She went on to explain to me that what some scientists do here is that they experiment with different characteristics of the mushroom material they grow. For example, I was able to touch and see one test product that had the texture and strength of wood. Another had the strength and texture of styrofoam--the very product that they set out to replace and vocally speak out against. In fact, if I hadn't been told otherwise, I would have easily believed that the product made from compostable eco-friendly mushroom material was styrofoam itself.

After I confirmed what I had to confirm with my mentor (regarding scheduling and the likes) my time at Ecovative had come to an end for the day but I knew that it was just the beginning. Even though I haven't actually started, I have a good feeling that I will come to love every minute of my time as an intern at this company.

Uses for Mushrooms Outside our Dinner Plate

Upon having found out that I have been paired with Ecovative, I was very excited and could not wait until I began. Typing in their website on the search bar, I was taken to their home page and there, I just started reading everything I could about the company.

Did you know that approximately 20 billion dollars of Styrofoam is created each year? It has been found that this very harmful product makes up 25% of our landfills and there they stay for thousands of years. According to Eben Bayer, the co-founder of Ecovative, a cube of Styrofoam is equivalent to a liter and a half of petrol. Thus, every year, each person is literally throwing away huge amounts of energy in the trash. Now if petrol, which is used to power almost everything around us, were unlimited, this maybe would not be that egregious of a crime. However, there is only a finite amount of petrol and this is where Ecovative fits in the big picture. It has been long known to everyone who has taken the required Bio-101 during their high-school years that fungi are decomposers that play a big role in the recycling process. Those at Ecovative have found and been able to use the vegetative part of the mushroom, mycelium, to create most of their products to-date. Due to the fact that mycelium takes things we would consider waste and transform them into an adhesive polymer, Ecovative has been able to use it as a glue to create all sorts of foam-like materials for packaging and insulation. Having started from what others thought was a crazy idea, founders Eben Bayer and Gavin McIntyre have truly started from the bottom and reached the top. Composed of with a team of engineers, scientists, product designers and more, Ecovative has worked and continues to work with an industry and consumers to “rid the world of toxic, unsustainable materials” for the betterment of not only the Earth, but also us humans as well.

reference: http://www.ted.com/talks/eben_bayer_are_mushrooms_the_new_plastic.html;
http://www.ecovativedesign.com/

The Beginning of the End: Senior Year Internship

Due to the fact that last year was such a rewarding experience in which I truly gained so much knowledge about working in labs, this year, I am all the more excited to partake in an internship. I really hope to learn many more skills that would help me in college—especially because I am looking into the science field. Last year, for the most part, I was given the task of calculations or in other words, figuring out how much of what solutions was needed. In addition, because of the fact that my particular internship (about DNA and RNA isolation) had many places for error, it was not altogether uncommon for the machine to malfunction and prevent my mentor and I from seeing the final results. Consequently, this year I would like to participate in experiments in which I get to do something from the start to the finish and then have the chance to thoroughly analyze the results. It would be all the more amazing if I could get the chance to form my own experiment and carry it straight through till the end. All in all, this year, I am simply just looking forward to further discovering and developing my lab expertise.

Looking Back

Looking back at what, I am really surprised to see how much I have grown as a scientist. Even before I first walked into RPI, I had great expectations. Because I wanted to pursue a carrer in the sciences, I was eager to get a first taste of what it would be like working in a white lab coat and preparing samples. In addition, because I was in AP Biology, I eager to put my new-found knowledge and vocabulary to use.

To my great delight, my internship proved to be just the thing I was looking for and starting with learning the chemistry of preparing solutes and substrates to preparing buffers and samples to learning about the workings of the machine to  learning how to interpret the final graph, I truly learned and attained a great deal about capillary electrophoresis as well as many useful lab skills. In particular, I still remember the feeling of contentment the day when I prepared buffers and DNA samples all by myself after receiving the desired measurements. Like that of a real scientist, I calculated and converted the required amounts and then assembled each respective solute via pipettes, scales, pH meters, etc... This said, weeks prior to that very day, I remember the great challenge of recalling my chemistry skills. Because of the fact that preparing samples and buffers all begin with the first step of figuring out how much of each substance to combine with another, I needed to have a solid understanding of chemistry before I started mixing and combining. Chemistry and math is not my strongest subject and because this was chemistry and math put together, I was all the more daunted. However, through this experience, I learned that the field of science is not just simply geared towards subjects like biology or chemistry or neuroscience, but that it is in more cases than not, a combination of them all.

For the incoming year, I would suggest having several more group meetings as I really liked hearing other interns talk about their experiences and the work they are involved with. Adding on to this, for the incoming new interns, I would like to advise that they should take the time to research more about their internship. (It might be helpful if they are told to do some research about their internship topics prior to their first meeting) Despite the fact that mentors explain in great depth the workings of the field, interns will come across times when even the explanations are too confusing to understand. In cases like this, I advise doing further research after at home, Emma, etc... In addition, I would like to tell future interns to never fear of asking their mentors too many questions--even the ones they think should be common sense! All in all, I find myself lucky to have been given this opportunity to further develop my interest and hone the particular type of career I want to pursue in the field of science and to have met such a great mentor.

My final poster summing up my vast knowledge about capillary electrophoresis ;)

05/01/13

May 1, 2013 was the last day for my internship regarding capillary electrophoresis. When I got to RPI, my mentor, rather than make another buffer or sample and run it through the capillary electrophoresis machine, she explained to me how to interpret even further graphs like the one attained a week before. As mentioned during my poster presentation (and probably somewhere in my previous blog posts), the DNA sample contains fluorescent markers that allow lasers to detect the DNA fragments. The graph produced at the end of the capillary electrophoresis process displays this test result and aids in interpreting the very fragments. The capillary machine determines the size of the DNA fragments in a sample based on the data detected by the lasers. The computer connected to the machine then depicts the lengths of the myriad of fragments as peaks on the graph. Thus, it helps determine the length of the detected DNA fragments.

The information accumulated all throughout several of these graphs is used to create the DNA profile. "Two points of reference are used to help the software as it determines the lengths: 1) the GeneScan software uses the internal size standard, which contains DNA fragments of known sizes; and 2) the Genotyper software uses allelic ladders as a point of comparison for the designation of the number of repeats in the DNA sample at particular chromosomal locations, since the peaks within the allelic ladder correspond to known fragment lengths at those locations."(An allelic ladder is a standard sample which contains all the alleles for a specific gene so that when it undergoes electrophoresis, there's a clear separation of the different alleles in the form of bands.)

The DNA Examiner works with both softwares and documents what the allele values are at each of the chromosomal locations analyzed and then compiles it to constitute a DNA profile (like the the graph of my previous post).

All in all, even to the very last day, just when I thought I knew all there was to capillary electrophoresis, I realized that my knowledge of it makes up only a parcel in the grand scheme of all things. ;)

04/24/13

At last! I got the results that I have been waiting for!

Today, I was able to see the final result from the electrophoresis machine. In the graph below, the peaks represent the corresponding 76-mers mentioned in my prior blog. (76-mers are single stranded DNAs that contain 76 basers). A while back, I prepared a similar sample for Yolanda with our end goal to find out the specific location of the particular 76mer within the prepared solution. Because this sample consists of a "spiking sample" (this increases the concentration of that particular 76mer in that particular tube so that when inserted into the machine and read, the graph will indicate the precise location with a peak that is higher than the other 9 polymers present in the mixture), my mentor and I were able to see that the spiked sample has 3 times the concentration of the other 76-mers--the highest peak being this very spiked peak.

In retrospection, I can't believe I have finally completed the whole process of capillary electrophoresis. Starting from learning about the chemistry of it all to actually preparing the buffers and samples, and then finally getting to use the capillary machine, I have learned so much from this experience. Next week, which I believe will be my final week, I am hoping we will be able to see more graphs like the one below and discover exciting results.

04/17/13

         Finally!! Today during my internship I was introduced/ got to use the capillary electrophoresis machine! Just because it was so long since I last wrote about the machine, I will give you a recap: In electrophoresis, proteins are separated and then characterized according to various characteristics (ie: size and charge) when moved across an applied electric field. Once completely separated, scientists are able to find abnormalities according to the types and densities of proteins in the sample. Consequently, they are then able to diagnose disease conditions. Unlike gel electrophoresis (in which the medium used for separation is a gel), capillary electrophoresis uses a liquid buffer <which now I am extremely good at preparing :) >. This gives the protein the ability to move more naturally and freely as compared to migrating through a gel. In short, capillary electrophoresis is a technique that separates ions with the use of an applied voltage.

 Today, I learned everything from how to start the machine, how and where to insert the buffers and DNA sample, how to run a test trial, and so on. Each side of the high volage supply is connected to an electrode which helps induce an electric field to initiate the movement of the sample from the anode to the cathode via the capillary tube. First, the capillary tube must be coated with the prepared buffer solution. "Electroosmotic flow is observed when an electric field is applied to a solution in a capillary that has fixed charges on its interior wall. When a buffer solution is placed inside the capillary, the charge is accumulated on the inner surface." Thus, before the sample DNA is inserted for reading, the capillary must be coated with the desired buffer.

     After the run through of the machine, using the buffer I prepared and the sample that my mentor pre-made, I got to use the machine. Putting everything together was not hard [this is one of the many advantages of capillary electrophoresis] as all I had to do was first rinse the tubes with water [cleaning it of remnants of the buffer solution from other runs] and then placing the two viles of buffer and sample in their respective places. I also made sure the temperature of the machine was what it needed to be as extreme temperatures could cause many malfunctions, ie: causing proteins to denature. Then, my mentor showed me how to take a reading of the sample via the hooked up computer. Because the explanation process concerning the workings of the machine and preparing another set of buffers took a fair amount of time, I did not get to see the produced graph (as it takes 30 minutes or more to produce 1 graph).

However, this coming Wednesday, rather than preparing the mixtures, I believe (and hope) I will get to spend more time using the capillary electrophoresis machine and learn how to interpret the resulting graph!

http://www.chem.sc.edu/faculty/morgan/lab.html

04/10/13

Today at my internship, I prepared yet another set of buffers. (I have made so many buffers now that it has become a procedure in which I can carry through while sleeping!) When last week, however, my buffers were created from solids (the various powdery substances), today, the six buffers I was told to prepare was created from liquids. The three were: CH₃COONa, NH₄HCO₃ and (NH)₂SO₄. I was told to measure out 0.4 mL and 0.2 mL of CH₃COONa and NH₄HCO₃ while measuring out 1.68 mL and 2.4 mL of (NH)₂SO₄. After this process, I diluted the solutes with distilled water to get a final volume of 6 mL for each of the six tubes I prepared. [all had a molarity of 0.5M]

            Once these steps were done, I quickly calibrated the pH meter using three beakers containing solutes with pH 7, 9 and 10 respectively. After this preliminary step was completed, I measured the pH levels of the six solvents I had concocted and labeled them accordingly. (In the end, the pH levels of all 6 tubes came about roughly to the estimated 8)

Today, like the week before, I spent the entire time consumed with preparing these buffers. In retrospection, I can definitely say that my science capability and knowledge—more specifically that my laboratory skills—has improved tremendously. My current capability to go in and prepare a buffer or a test solvent based on measurements (numbers) alone never fails to amaze me. Next week, I am hoping to be doing something other than preparing buffers and actually moving towards working with the capillary electrophoresis machine.