It costs a lot. Maybe about $2M?

The average biotech lab has at LEAST a micro, analytical and top loader balances, ultrasonicator bath, HPLC with varying detectors (UV, PDA, MS), HPLC columns, a chromatography aquisition server(ie Empower), validated sample storage chambers at all the ICH conditions (5C, 25C/60%RH, 30C/65% RH, 40C/75%RH), Karl Fisher, Fume hood, lab benches, Wet and dry chemicals, various glassware (pipettes, beakers, volumetric flasks, graduated cylinders etc), Dissolution bath if you are planning on doing oral dosage units (inhaled pharmaceutical and sterile injectables require a lot of other specialized equipment), a stand alone UV-Vis, and maybe a GC. In addition to gathering chemicals for a home chemistry kit, be sure to collect a few everyday materials, too. measuring cup A glass measuring cup is useful for many activities. measuring cups/spoons Don't worry. You can still use them for baking and other cooking. Just be sure you have a way to measure both liquids and solids. coffee filters Coffee filters make great covers for projects that require evaporation, but not dust and bugs. They make nice pH paper and chromatography paper, plus they are good for their designed purpose, filtering. clear glass jars Most of the time, you can use dishes from the kitchen for projects, but sometimes you'll want a clear container you can throw away when a project is completed. string String is used for growing crystals. Nylon line is the best for large single crystals. Cotton string is good for masses of crystals. resealable bags The quart size is probably useful for the most projects. paper towels Chemistry isn't always neat and clean. Projects that involve food coloring could stain cloth towels. Cheap PCR: new low cost machines challenge traditional designs Despite price drops, thermal cyclers still remain in the exclusive domain of the research laboratory. Andrew Wiecek talks with three groups working to change this by lowering PCR costs in an attempt to bring this key technology to everyone. Although PCR is a fundamental tool for many biologists, thermal cyclersessential to driving PCR reactionshave not yet become easily affordable or widely accessible. An instruments expense, which is between $4000 and $10,000, has essentially confined its usage to university laboratories, and excluded entities like public schools or field-based health initiatives, which could benefit from easy access to DNA amplification technologies. Cheaper thermal cyclers can be found through online auction sites such as eBay for as little as $300, but these machines are often sold as-is, and lack the conveniences of newer devices. Half the time it doesnt work right, says Josh Perfetto of OpenPCR, a group currently developing low-cost PCR machines. If you do get a working one, its typically an older unit. It doesnt have fast ramp time rates. It doesnt have a heated lid. While researchers experienced in PCR can work without these features, it creates another barrier for less-skilled users. Anything we could do with the hardware to make it more robust, I think thats good, says Perfetto. For example, I think its important for people to have a heated lid rather than putting oil on the PCR reaction. It creates another step that they can screw up or contaminate. But a change might be on the horizon. Three companies have been trying to develop affordable, robust open-source PCR machines. Working out of bedrooms and borrowed lab space, each groups design choice reflects its particular aspirations, but they all have the same ultimate goal in mindto lower the financial and knowledge barriers and bring PCR to the world at large. Polymerase education

Otyp's prototype for their thermal cycler, which they plan to lease to schools for educational purposes. Source: James Peyer, Otyp James Peyer knew he had to develop an open-source PCR machine. A first-year doctoral student at the University of Michigan studying stem cell biology, the 23-year-old is interested in advancing high school science education. His idea is to develop laboratory exercises in which students perform advanced experiments such as genetically engineeringEscherichia coli to fluoresce green. But such experiments require PCR machines and pipets, which most schools cannot afford.

With the help of his younger brother David, a 20-year-old biomedical engineering undergraduate at the Miami University of Ohio, Peyer founded Otyp, a company that leases PCR machines and pipets to schools. Their goal is to provide kits containing leased equipment and necessary reagents for $400, which would fit most schools budgets. But Peyer quickly found he could not find reliable PCR machines at this price. Even with bulk discounts, I was still paying about $2000 for a PCR machine, which meant that even when I boiled down the cost as much as possible it still cost a school about $600, says Peyer. To avoid an unsustainable business model; Otyp trimmed its margins and currently offers the educational kit to schools for that price. Schools dont have the money to spend $600 on a 2-week laboratory course, even though its important, says Peyer. He needed to come up with a cheaper way of acquiring working PCR machines, or Otyps educational leasing kits would only be affordable to a limited number of schools. With no other option, Peyer and his colleagues decided to build their own instrument. Considering the current PCR marketplace, such a reinvention seems bold for a group of twentysomethings. But Peyer keeps his mind on the basics. A PCR machine is pretty simple; its just an aluminum block that heats and cools on a schedule, he notes. With the help of the newest member of their team, Kyle Lawson, a 23-year-old industrial design graduate from Savannah College of Art and Design, the Peyer brothers spent the last four months working in a makeshift laboratory set-up in Peyers apartment bedroom. Keep it simple and straightforward is the design principle for Otyps PCR machine prototype, and the team has used open-source protocols to cut costs and increase accessibility. Our main goal was to cut out the fat of existing PCR machines, says Peyer. Instead of using an injection-molded plastic case requiring expensive custom models and special plastic to withstand high heat , Peyers group went with a simple steel computer case. And rather than using an integrated computer with touch-screen controls and custom circuitry controlled by a proprietary operating system, the Otyp PCR machine will be controlled by the Arduino open source microcontroller and a standard PC through a USB port. Trying to figure out what circuitry we want to use and how to write the program to control the machine was a pretty long and uphill process that were still grappling with, says Peyer, who has no programming or electrical engineering background. The prototype costs $300, not including labor, and the group hopes to refine its design during next year in order to decide whether to go into large-scale production in early 2012. It will help us reduce the costs from $600 per school to $400, and if we can do that, thats only a hair over what they are already spending for gene transfer kits [from other science education suppliers], says Peyer. These kits eliminate the amplification step and bypass the need for a PCR machine. If we can get the costs down to that level, then well be able to go viral as far as the education market is concerned. Groovy amplification

The first LavaAmp prototype weighs only 180 grams, fits in the palm of a hand, and is powered by a USB cable or 4 AA batteries. Source: Jim Hardy, LavaAmp Outbreaks of Chagas disease are on the rise, affecting about 10 million people living in endemic Latin American countries. The disease is caused by a parasite, and leads to swelling and potentially fatal heart and digestive system disorders in chronic cases. There are people that suffer but have no way to know if they are affected, says Guido Nez-Mujica, a Venzuelan computational biologist. Chagas outbreaks in his home country inspired Nez-Mujica to find a cheap, portable PCR machine to help diagnose those infected with Chagas and other neglected diseases in developing countries. Portable detection devices are necessary in third-world countries because those infected often live in rural areas. Chagas, for example, is transmitted by triatomine bugs, which live in the cracks of homes in rural or suburban communities. Treating these individuals can be a multiple-day journey: a doctor must travel to the patient to take a sample, return to a laboratory in an urban area, and then return to the patient to present the results and begin treatment. However, with a portable PCR device, diagnosis and care can begin immediately. At the 2008 SciFoo campa weekend retreat for scientists, technologists, and writers, and organized by Google andNatureNez-Mujica found four kindred spirits: open science philosopher Joseph Jackson, former Life Technologies researcher and biotechnology entrepreneur Jim Hardy, and engineers Rik Wehbring and Rob Carlson of Biodesic LLC (Seattle, WA). After the conference, they founded LavaAmp to develop a new breed of portable, cheap PCR machines. Make the device rather inexpensive, make it portable, make it run on batteries if you need to, and make it lightweight. Thats the device that were working on now, says Hardy. Their first prototype weighs only 180 grams, fits in the palm of a hand, and is powered by a USB cable or 4 AA batteries. The LavaAmp team reduced the overall size by eliminating a standard part of typical PCR machines: the aluminum block, which is used to heat and cool the reactions Its a buoyancy-driven convection current of different thermal zones created by different heating elements, says Hardy. And the liquid circulates through different zones. Thats how we get the PCR.

While most PCR machines are designed for high-throughput, the LavaAmp instrument takes the opposite approach. You dont have the need to run 96 samples at once, says Hardy. For most applications, thats not necessary. While the industry-standard 96- and 384-well formats are really good for screening, says Hardy, its not practical for diagnostics purposes. Rather than this high-throughput approach, the LavaAmp PCR device can run 2025 samples at once. The LavaAmps thermal-gradient convection currents provide three different temperature zones that perform denaturation, annealing, and extension. The platform itself made it much simpler, says Hardy. Theres not a need for a lot of sophisticated electronics. The biggest technical issue that remains is a convenient way of loading and unloading the samples into the systems loops. The loops have a very small volume, says Hardy. Trying to keep air bubbles out is a problem because air bubbles disrupt the current. While the initial price will be about $300 to $500, the groups ultimate goal is to design an instrument that will cost less than $100. The device is just going to be more or less disposable. You can use it two or three times and just get another one if you leave it behind somewhere, says Hardy. The first commercial versions are expected to be available in early 2011. Hack this PCR!

Although the OpenPCR machine will have a rather traditional design with an aluminum block, it will have an untraditional case made from birch wood. Source: Josh Perfetto, OpenPCR At the January 2010 Outlaw Biology conference, Perfetto and software developer Tito Jankowski heard first-hand from several individuals about how the DIY Bio community could benefit from a cheap, open-source thermal cycler that could be adapted for different applications. Driving back from the conference, the pair agreed that they could fill this void. Thus, the OpenPCR project began. According to Perfetto, OpenPCR started more as an open hardware project, not so much as a big company start-up. But as weve gone forward, new ideas have emerged and we think we could meet some potentially bigger things coming out of this, he adds. Neither Perfetto nor Jankowski has formal training in biology, but they have been involved in the California DIY Bio community. Jankowski was part of the Brown University iGEM team that designed an open source gel electrophoresis box, which is now being sold by Pearl Biotech for $499. Perfetto is excited about the potential of the DIY Bio community. Ive worked on a lot of mobile phone software and rep software, says Perfetto. Theres still innovation, but the field is relatively mature. Biotech is where were going to see a lot more innovation and excitement for the next coming years. Although the OpenPCR machine will have a rather traditional design with an aluminum block, Perfetto and Jankowski have experimented with different parts and materials to create the fastest, cheapest machine for the unskilled biology enthusiast. The machines design features a heated lid, which is essential, according to Perfetto, because untrained users struggle with the mineral oil addition step to reduce condensation. The group considered different heating and cooling optionsincluding convection and radiation to cool the heat blockbut they ultimately decided on a complex Peltier cooling solution to quicken experiments and enable the use of cheaper reagents. And, instead of permanently gluing the case shut, the case will be shut with bolts for easy removal for their bio-hacker clients. In two months, OpenPCR kits will be available for biology hobbyists for $300400, and will include all the necessary parts and schematics. Pre-assembled machines will also be offered as well. While OpenPCR is initially targeting the hobby and educational markets, Perfetto and Jankowski do have some nextgeneration ideas, including developing a line of customized detection devices that could be used by anyone, even those without an understanding of PCR. Were focusing on the user interface and making it easier, says Perfetto. We want enable people that dont have a strong biotech background and dont know the details of PCR to use this device to answer questions that interest them. PCR for all Cheaper PCR machines are well on their on their way in their promise of educating the next generation of scientists, diagnosing diseases more quickly in third-world countries, and changing PCR applications around the globe. Although each effort is separate, Jackson from LavaAmp explains that it all stems from a small but growing community attempting to push open-source biotechnology innovation. They all know each other, he says. So it looks like there is this demand, and at least were converging. He says the next year, when these cheap machines begin stocking shelves, will indicate how people want to use them. So how many people are going to be running PCR in their kitchen? Were all interested in seeing the applications of the machines. MacGyver Extraction of DNA: Reagents and materials for this portion of the experiment are fairly straightforward. Here, one needs to find a tissue source, some manner of physical breaking, clean water/buffer solution, a soap, and an alcohol. In addition, some type of container to do this in will be required, preferably one that is transparent in nature.

Tissue Source: Although the chemical characteristics of the DNA isolated is practically identical from organism to organism, the cell in which it is housed can vary greatly. Consequently, choice of tissue is the largest variable, but arguably the best element for a student to play around with. We have tried extracting DNA from onion, split peas, corn, yeast, bean sprouts, wheat germ, kiwi, banana and even human cheek cells, all with varying degrees of success. We find that overall, good DNA can be achieved with any fresh produce or grain material, as long as the experimenter is willing to play around to find optimal conditions. Overall, however, we recommend onion or banana tissue as the best sources of low maintenance DNA extraction. We also recommend wheat germ, in that a fast procedure exists that works very well. Finally, DNA isolated from a students own cheek cells is an interesting alternative, since it brings in a personal element to the activity. However, it should be noted that isolation from cheek cells is less reliable. Physical Step: Depending on the tissue you use, a physical step may be warranted. This could be as involved as cutting up the tissue and then using a mortar and pestle, or as minor as banging around with a wooden chopstick. In general, plant material and meat cuts would benefit from such a step, since plants have a tough cell wall to contend with, and meat usually contains tough muscle striations. Buffer Solution: This is the fluid that the sample needs to be immersed in, and as such has the role of simply keeping your DNA safe. There are many possible MacGyver type buffers that one can use for the extraction procedure, which all basically work (see Table 1). We should note, however, that distilled or bottled water should be used. Tap water appears to have a minor effect on the extraction procedure, and is actually very detrimental in the subsequent gel steps described later. TABLE 1: BUFFER RECIPES* A: Saline Buffer 0.9% saline (contact lens solution) B: Regular Buffer 1.5g table salt (NaCl) 5.0g baking soda (sodium bicarbonate) to a final volume of 120ml with water C: Acidic Buffer 1.5g table salt (NaCl) 5.0g baking soda (sodium bicarbonate) 5mL of vinegar to a final volume of 120ml with water D: Proteinase Buffer 1.5g table salt 5.0g baking soda 2.5mL Complete eye solution or 2 protein tablets (Complete brand) to a final volume of 120ml with water E: Meat Tenderizer Buffer 100mL hot water 3g meat tenderizer *Note that water alone can also be used effectively. Soap/Detergent: Liquid dish washing soap was generally used. In terms of specifics, we found better results when the soap was colourless, and found that unscented versions may work best overall. Essentially, most soaps available at grocery stores have additives that play roles in preservation, scent, colour, stain removal etc, etc. Although these things are usually not an issue with the technique, choosing the simplest soap is a good thing to troubleshoot with if you are not having success with your extraction. In general, we found that using about 5ml (or less) of detergent per 120ml solution works well. Filtering: Depending on your tissue choice and also the amount of tissue you have decided to use, it may be pertinent to include a filtering step to remove the debris. This allows a better visualization of the DNA snot effect at the end. Filtering can be easily accomplished using coffee filters, or cheese cloth. Note that when using cheese cloth, you will need use 3-4 layers of cheesecloth. Coffee filters are generally easier because they are cheaper, more accessible, and easier to cleanup. Precipitation with Alcohol: Precipitation of DNA is done by contact with alcohol. This alcohol is available at pharmacies as rubbing alcohol of the 99% isopropanol variety. Alternatively, most schools will have access to 95% ethanol stocks. The alcohol should be added in a layering manner so that one can see the DNA forming at the interface between your sample and the alcohol. You should add at least two times the volume of your original sample. If you do not see snot forming, then mixing everything together should get the desired effect. Note that if you intend to resort to mixing, then the filtering step may be especially necessary for ease of visualization. DNA precipitates under alcohol treatment because it is naturally hydrophobic and as such will tend to clump together if the solvent is not optimal (i.e. water). As a result, adding an alcohol to your prep will mean that DNA is not completely solvated in its optimal environment (water), and will therefore aggregate, and precipitate out of solution. It should be stressed that the degree and ease of clumping will depend on the amount of DNA present, and also the concentration at which it exists. Therefore, if you see minimal amounts of DNA, you should be able to correct this by (i) trying to isolate from more tissue, and (ii) resuspend the material in a smaller volume of fluid. TABLE 2: SUGGESTED MACGYVER GENOMIC DNA EXTRACTION PROTOCOLS: GENERAL PROTOCOL 1. If necessary, slice up DNA source of choice. Use an amount about the size of a strawberry. 2. Using a mortar and pestle, grind up sample while gradually adding 10mL of prepared buffer solution with the detergent already added. Grind for at least 5 minutes with all of your weight and strength to ensure that you break open the cell membrane and reach a

creamy soup consistency. If the sample is too thick after grinding, add more saline solution to achieve the optimal thickness so that the liquid portion of the sample is able to pass through the filter, while the larger cellular material remains behind on the filter. Note: If the DNA sample is frozen, it is considerably easier to grind. 3. Filter your samples juice into a small beaker. Let the solution drip into the beaker until all of the liquid has passed through the filter. If this takes too long, simply squeeze all of the juice from the sample through the filter. 4. Add 2 volumes (this means approximately two times the volume of the sample present) of ice cold alcohol down the side of the beaker with a straw or pasteur pipette. Do this step slowly to enable the alcohol to form a layer on top of the juice layer. As you let the beaker sit, the DNA should precipitate. The longer you wait, the more DNA you should see. If you dont see precipitation, gently mix everything together. 5. The DNA precipitates should resemble a thread of translucent white snot,at the interface between the juice and alcohol. After a considerable amount of time, the DNA may eventually float to the top of the alcohol layer. 6. You can remove the DNA with a wooden popsicle stick or glass rod. DNA adheres well to the wood. QUICK PROTOCOL 1. Place a teaspoon or less of wheat germ in your cup. 2. Add about 10ml distilled water and crush gently with a popsicle stick/chopstick for 1 minute. 3. Then add a squirt of dish detergent and crush gently with popsicle stick for 2 minutes. 4. Slowly pour alcohol down the length of the stir stick, layering the alcohol on top of the water. 5. Set the container down on a table and look for the DNA at the interface between the alcohol and water. CHEEK CELL PROTOCOL 1. Measure 10 ml of regular buffer (from Table 1), pour buffer in mouth and swirl around cheeks for about 1 minute. 2. Spit the water back into a container, preferably something relatively narrow like a test tube. 3. Squirt a bit of liquid soap to the sample and mix well with popsicle stick. If you can mix by inversion, then do so gently about 20 times. 4. Add 10 ml cold alcohol slowly to the sample and make sure to pour it at an angle down the side of the test tube so that two layers are formed. Do this very gently, with a straw, etc. Its important that the two layers are not disturbed. 5. Wait for about 10 minutes and the DNA will appear afloat on the alcohol layer. Gel Electrophoresis of DNA in a Research Setting: Electrophoresis is a way of separating molecules based on charge and size. In our case, we want to separate different sizes of genomic DNA molecules obtained from fruits, vegetables and yeast. Generally, polysaccharide polymers such as agarose or acrylamide are used to form the electrophoresis gels. Because DNA is negatively charged, one can force it to travel through the gel by applying an electric field in the system. Normally, this is achieved by using special gel apparatus designed to facilitate the production or casting of the gel as well as allow a platform to immerse the gel in an ion containing buffer to create an electric field. Such an apparatus will run a minimum of about $500, and power packs normally used to deliver ~80V of voltage can run in a similar price range. Consequently, this aspect of the MacGyver project is arguably focused on cost savings and concentrates primarily on finding a substitute for agarose and directions for producing a gel apparatus. MacGyver Gel Electrophoresis of DNA: Gel Material: Agarose is a component of seaweed and as such is a refined purified molecule derived from a common food thickener known as Agar Agar. which incidentally can be easily found at most oriental style food stores. Agar Agar can be purchased in either a flake, noodle or powder form. Try to ensure that it does not contain any additional ingredients (such as glucose) as these ingredients may interfere with the formation of the gel matrix. To make the gel, it is recommended to use Agar Agar in the powder form rather than the flake or noodle form. The other larger forms tend to require cutting and additional filtering which is problematic and very messy. Running Buffer: You will need to prepare a running buffer which is required to make the gel, and also required as the fluid that will ultimately immerse your solidified gel to allow the electric field to be conducted. The Macgyver running buffer recipe is as follows: - 0.05g of NaCl (this is the principle ion) - 2g of Baking Soda (Sodium Bicarbonate) - bring to 1L with distilled bottle water pHd using pet store aquarium pH kit to approximately pH7.5. (we used alkaline buffer made by Seqchem). Gel Apparatus: Although a research grade gel box is costly, it is in fact a relatively simple piece of equipment. In essence it is a large container (lets call it a buffer chamber) that can hold fluid, whereby opposite ends are connected to a power source setting up a positive/negative electrode scenario. The buffer chamber needs to be able to conveniently hold a smaller container (lets call this the gel casting chamber) that is used to make and hold the gel. Furthermore, the smaller gel casting chamber needs to fit in such a manner as to be in the middle between the two electrodes of the buffer chamber. Assembling this electrophoresis box should be quite straightforward and even enjoyable for those that like to make things. We have included here a cartoon guide (Figure 1) for making these two chambers out of common plasticware (tupperware, soap dish, etc). For the electrode connections, we were limited to stainless steel screws (5cm) and stainless steel wire (20 gauge), which worked fine. However, a more elaborate electrode system would be easy to make with a visit to a proper hardware store. As an alternative to using tupperware, we have also found Lego to be extremely useful in custom making a buffer chamber to fit your gel casting chamber. Note that Lego chambers will need to be lined with a water tight material, but recently Glad has developed a new Glad PressN Seal material which works perfectly and is easy to handle. Finally, a comb: will need to be made. This is a contraption that allows small wells to be formed in your gel. Here, we found lego to be especially useful, but in a bind, we also made combs by cutting out pieces of plastic, or taping the teeth of a real hair comb. FIGURE 1: MACGYVER GEL BOX CONSTRUCTION NOTES.

Gel Preparation: Using the powdered Agar Agar you will want to make a 1.2% to 1.5% gel (w/v or 1.2g to 1.5g per 100ml of running buffer). In a separate container, (a flask for instance) weigh out the required amount of Agar Agar and add the required amount of running buffer (specific amounts will be dictated by the size of your gel casting set-up). The flask should be large enough so that the Agar Agar solution forms a 2 cm layer at the base. This is required to ensure a large surface area in contact with the hotplate for effective melting. It is recommended to use a hot plate and continued stirring to heat the agar solution evenly. An alternative is to use a microwave, but with this method one must use a low heat level at short intervals while swirling the mixture frequently. If this procedure is not carried out carefully, the solution will likely overboil and create a large mess. Note that if the Agar Agar solution is not completely dissolved, this will greatly affect the mobility of the DNA as it travels through the gel. Furthermore, undissolved bits of agar will be stained during the staining procedure, making it difficult to view the bands of DNA. Note: some schools have access to the Agar used to make bacterial plates. This material works very well in pouring gels (much better than Agar Agar) We recommend making a 1% w/v gel for genomic visualization purposes. Make sure that you have sealed the open ends of your casting gel chamber with masking tape or similar material. Pour the melted mixture into your casting gel chamber. Remember to also insert the comb so that well formation can occur. It will take approximately 20 to 30 minutes at room temperature for the gel to solidify. During this time do not disturb the gel. The gel is quite delicate, so be very careful when you remove the comb before loading your sample The gel should be poured to about a 0.5cm to 1.0cm thickness. Thicker gels allow the opportunity to make deeper wells, thereby allowing a larger sample to be loaded. Thinner gels should run faster. Note that if the gel is too thin, it may float when immersed. Try to use the gel as soon as possible. Although you can wrap it in plastic wrap and store it in the fridge overnight, they will work better when used fresh. DNA Sample Preparation: Your genomic snot DNA can be removed at the end of the extraction procedure using a wooden stir stick. The DNA should then be dipped in a 70% alcohol solution which facilitates removal of excess salts. This is very important for effective dissolving as well as effective gel running. The DNA can then be air dried for approximately 10 minutes to evaporate residual alcohol, and then dislodged into a small volume of running buffer (the smaller the better, i.e. <0.5mls). One should note that genomic DNA can often take days to dissolve properly. We suggest allowing the DNA to dissolve overnight at room temperature (if you have access to temperature up to 50C, then that is best). Be gentle to your sample as the large genomic DNA is very prone to shearing. Once the overnight dissolving step is finished, consider this your DNA sample regardless of whether it has dissolved to completion. Your sample will then need to be treated with a loading buffer. This is essentially something that will cause your sample to be viscous so that it can indeed sink into your wells during loading. Often, a loading buffer also has a dye, which will travel in the same direction as the DNA, and also makes the gel loading easier to see. Our recipe is as follows: - 0.5ml glycerol/glycerine (available at the pharmacy section) - 0.1ml distilled water - several drops of Club House Red Food Colouring (note that choice of dye can affect outcome greatly we didnt have a lot of success finding something good here, so a last resort would be to use without a dye). This loading buffer can be used as 5X to 10X meaning that for 0.5ml of sample, you would need to add 0.05 to 0.1ml of loading buffer. Mix carefully. Your sample is now ready for the gel. Gel Running. Place your solidified gel (in gel casting chamber) within the larger buffer chamber. Add running buffer until the gel is immersed such that there is at least 3mm of fluid above the gel. Keep in mind that the more fluid you add, the slower the gel will run. To your wells, load as much sample (with loading buffer) as possible. This is probably best done with a plastic stir stick attached to some type of rubber bulb. Essentially you would like to assemble something that can deliver fluid through a small tube. Once all the samples have been loaded, you will want to connect the electrodes to a series of 9V batteries. Your DNA is negatively charged so you

want to position the positive electrode at the end away from the wells. Basically, one battery will suffice but will be very very slow (overnight run scenario). 5 to 7 or them lined up in a series circuit should deliver a good amount of voltage. Note that when the gel is running, DO NOT stick your finger in the fluid. Depending on the number of batteries you use, as much as 100mAmps of current is delivered (enough to give you a small shock). When the circuit is running, a good visual check is to see that bubbles are forming from the wire electrodes, and usually most visible at the positive end. The optimal amount of time to run the gel is, frankly, something that is difficult to predict, as it depends on the size of your gel, the thickness of your gel, the amount of running buffer in the system, the amount of voltage applied, and even the wiring set-up used. Consequently, this is the one thing where you will definitely have to play around. However, with a 5 to 7 battery set-up, you will need at least a minimum of 1hour, and possibly more. In addition, if differentiating genomic preps is in order (i.e. different genome size), you may need to experiment further with run time to see this potential difference. Visualizing DNA in a Research Setting: Upon completion of electrophoresis, the location of the bands need to be visualized. A common way to detect the bands is to stain the gel with ethidium bromide, which fluoresces under ultraviolet light to view the DNA. Unfortunately, both ethidium bromide and ultraviolet light are hazardous (Ethidium is higly carcinogenic and UV light can burn a persons eyes without proper safety measures) and are therefore, not ideal for use among high school students. MacGyver Way to Visualize the DNA: To visualize the DNA bands, the gel was stained in a Methylene Blue solution. Methylene Blue consists of the salt methylene blue chloride. In water, the salt disassociates into a positively charged methylene blue ion that is colored blue and a negatively charged chloride ion, which is colorless. This blue chromophore is then able to bind to the positively charged DNA in the gel. Methylene blue is a convenient stain to use in the lab because it is chemically safe, reusable, and detects the presence of more than 20ng DNA/band. More importantly, methylene blue can conveniently be found at most pet stores. It is generally sold as an aquarium disinfectant at a 5% concentration. The best gel staining results were obtained immersing the finished gel in a 0.02% methylene blue (in distilled water) solution overnight at room temperature. Using this protocol, destaining excess the excess colour with distilled water incubations was not required. We found that higher concentration solutions tended to stain the gel too dark making it difficult to differentiate the background from the DNA bands. This was observed even after destaining with distilled water. Also, if the Agar Agar powder was not completely dissolved in the gel, the non-dissolved flakes were stained dark blue, preventing the formation of distinct, visible bands. FIGURE 2: CORN GENOMIC DNA PREP MACGYVER STYLE. Yes, its faint! But its there

Conclusions: Overall, we have described an effective outline of methods to perform some basic molecular biology techniques under the MacGyver limitation (Figure 2). We should stress however that doing this in your own classroom will inevitably require some working out of your own. This is due to a multitude of considerations such as Do I have enough DNA? what is my gel set-up what type of specialized reagents do I have at my disposal to make things even more efficient? etc. However, it is hoped that the information presented here will make your troubleshooting as smooth as possible. Good luck!

Electrophoresis is a method used with agarose gels to separate different size strands of DNA, consisting of genomic
DNA and plasmids, or the products of restriction enzyme digests and PCR. The following is a simple step-by-step procedure for running a gel and visualizing DNA fragments by ethidium bromide staining. Time Required: Approximately 2 hours Here's How: 1. Choose an Electrophoresis Buffer Before you prepare the molten gel, you need to decide which electrophoresis buffer to use. Once that is decided, mix the powder agarose with the buffer at the desired concentration, depending on the size range of the DNA strands you are working with. The gel is melted by heating, often in a microwave. The dye for visualizing the separated DNA can also be added at this stage, if you want it within the gel. Alternatively, the gel can be stained by soaking it in dye solution after it is run. 2. Pour the gel. Prepare the gel casting tray by taping or blocking the ends as indicated by the manufacturer. Add the molten agarose to the casting tray and let the gel set. Wait until the gel has cooled to approximately 60 degrees Celsius before pouring, to avoid warping the plastic of the casting tray. Remember to place the sample combs in the gel casting tray prior to pouring the agarose. The combs form wells in the gel into which the samples are deposited prior to running the gel. Allow the gel to cool at room temperature, or, if in a rush, in the refrigerator. 3. Prepare the gel apparatus Remove the sample combs. Carefully remove the combs by pulling straight up out of the gel. Place the gel, still in the casting tray, into the electrophoresis chamber. Remember to remove the end pieces of the casting tray, or tape in some cases, at either end of the gel. Place the casting tray in the gel apparatus such that the end with the wells and the opposite end are in contact with the loading buffer. Also ensure the end with the wells is in line with the cathode, which is usually colored black. The DNA is negatively charged and will migrate towards the positively charged (red) anode.

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Prepare Samples DNA samples are mixed with loading buffer usually at a ratio of about 1:5 or 1:10. The loading buffer contains a dense substance, usually glycerol, that causes samples to sink to the bottom of the wells, preventing them from diffusing into the running buffer. Tracking dyes present in the loading buffer migrate along with the DNA, ideally with a separation that leaves one migrating behind, and one migrating ahead of the sample. Bromophenol blue and xylene cyanol dyes are often used and migrate at similar rates to double-stranded DNA fragments of 300 and 4000 base-pairs, respectively. Load the Gel Transfer the DNA samples into the wells of the gel using a pipette. Loading DNA samples must be done slowly and smoothly to prevent sample from squirting up into the running buffer and diffusing away. Run the Gel Place the lid on the gel apparatus and connect the leads to a power box. Apply current by turning on the power box and watch for bubbles to form, confirming that a current has been applied. Watch for the tracking dyes to begin migrating in the proper direction (toward the other end of the gel), confirming that the gel has been placed in the chamber with the correct orientation. Generally gels are run at approximately 80 - 100 mAmp. A setting any higher will cause the buffer to heat up to the point that it begins to melt the gel. Monitor the Tracking Dyes Watch the progress of the tracking dyes and turn off the current before the leading dye has reached the opposite end of the gel. Stain the Gel If dye was not added to the molten gel, stain the gel now in an aqueous solution at concentrations suggested by the supplier. For example, ethidium bromide is used at about 0.5 ug/mL and stained for about 15 to 30 minutes, then destained or rinsed in water, for 10-15 minutes. Some other dyes are available that fluoresce more strongly and might not require destaining. Most are fluorescent dyes that intercalate between the bases of DNA and RNA. Be careful handling them, as this property also makes them potentially very potent carcinogens. Visualize the DNA Place the gel, either alone or in the casting tray, on the transilluminator (UV light of wavelength 254 nm). Remember to wear protective eyewear. Take a photo, if desired, and if you have the equipment. Visualization and photography should be performed shortly after running the gel, as DNA will diffuse over time and the bands will become blurry.

Load the gel slowly using a small pipette tip, to avoid agitating the running buffer and losing sample from the wells. 2. Always wear gloves when handling ethidium bromide and remember protective eyewear when using the transilluminator. What You Need An electrophoresis chamber and power supply. Gel casting trays composed of UV-transparent plastic. + Sample "combs". Electrophoresis buffer (usually TAE or TBE). Loading buffer containing tracking dyes. Ethidium bromide solution. Transilluminator (UV light box) and photography equipment (optional). How To Increase IQ? Working Memory Training, Smart Drugs and tDCS Reviewed 3 May 2013 | 5 Comments | posted by Mark A. Smith | in Biohacking, Brain Training & Mindware, Increase IQ,Mindhacking, N-Back Training, Self Quantification, Smart Drugs, tDCS, Working Memory Training IQ Defined Our intelligence measured by valid IQ tests is our ability to grasp situations, reason, problem solve, and learn and act efficiently and effectively. David Wechsler the creator of the most widely used IQ test, the WAIS defined intelligence as: the global capacity of the individual to act purposefully, to think rationally and to deal effectively with his environment. Intelligence is better conceived as being switched on and competent rather than being just book smart or good at math. As Napoleon Hill put it: Action is the real measure of intelligence. The value of IQ IQ level is known to be positively correlated with many valuable things. Some that have been demonstrated in peer reviewed research are: achievement motivation, altruism, artistic ability, creativity, dietary preference, educational attainment, emotional sensitivity, health, sense of humor, income, breadth and depth of interests, leadership, longevity, linguistic abilities, memory, moral reasoning, motor skills, occupational status and success, and social skills. IQ is inversely associated with accident proneness, obedience, alcoholism, authoritarianism, crime, dogmatism, neurosis, impulsivity, racial prejudice, smoking and obesity. The practical advantage of having a high IQ increases as our work/career environments become more changeable and complex more novel, ambiguous, unpredictable, or multifaceted. A high IQ is key to strategic thinking in which planning, decision making and problem solving unfolds in the midst of complexity and uncertainty. IQ is thus of prime value for entrepreneurs who are strategizing and problem solving their way to success in far from stable environments. The more new situations you experience, the greater your ability to adapt to ever-changing circumstances. For a longterm employee, being laid off may come as a serious blow. But for a long-term entrepreneur, losing a particular client is

Tips: 1.

just par for the course. The entrepreneur has learned [how to] make it easy to add new income streams, while the employee may have much lower intelligence in this area. Similarly, people who interact socially with new people every day will develop much greater social intelligence than those who interact with the same people over and over. Steve Pavlina IQ Increasing Technologies: A Review This article reviews three of the most effective IQ-increasing interventions that have a firm scientific basis a basis in experimental laboratories and the exacting standards of peer reviewed scientific journals. The methods described below are part of the accumulated understanding of the scientific community about what can increase IQ not just temporarily but long-term. Cognitive-enhancing nutrition, exercise and meditation is not covered in this review, which focuses on the use of intervention technologies. 1. Brain Training Software Far-reaching advances in cognitive psychology and cognitive neuroscience over the past decade have identified a close link between frontal lobe working memory circuitry, and fronto-parietal problem solving, self-control and fluid reasoning circuitry. Our working memory is used for holding information in mind (images, concepts, language, numbers) for brief periods while engaging in active, goal-focused thinking or comprehension, while screening out distracting information. Working memory has a limited capacity, and the bigger that capacity the more the cognitive RAM power a person has for processing information to make connections, generate alternatives, and grasp relationships. This brainpower lies at the core of being smart. If super brain Eddie Morra in Limitless changed one thing in his brain, it was his working memory circuitry!

Software has now been developed for selectively targeting working memory circuitry, resulting in long term neuroplasticity changes increasing short term memory capacity, problem solving ability, self-control and overall IQ. This software is based on a training exercise called the n-back. A scientifically credible product is found at IQMindware. A review published this year on the effectiveness of n-back working memory training by an old grad school friend, Dr Jason Chein, concludes: core working memory training studies seem to produce far-reaching transfer effects, likely because they target domaingeneral mechanisms of working memory. The results of individual studies encourage optimism regarding the value of working memory training as a tool for general cognitive enhancement. (Quoted from: Does working memory training work? Psychon Bull Rev 2011) In choosing an n-back working memory training application, ensure that you have a version and training program that has been demonstrated to increase IQ in a published scientific study such as the landmark paper by Dr. Susan Jaeggi and colleagues (link) that first drew public attention to the benefits of n-back training. There are a number of n-back training programs on the market that do not replicate what is known to work. Avoid them. 2. Nootropics (Smart Drugs) The issue of using medication for cognitive enhancement is highly controversial, but the ethics of smart drugs is not discussed in this article. Im simply presenting the facts. Nootropics also known as smart drugs, memory enhancers, cognitive enhancers and intelligence enhancers are drugs, supplements, nutraceuticals (a product isolated or purified from foods) that are designed to improve cognitive functions such as memory, attention and intelligence. The use of nootropics for cognitive performance is widespread. In January, the prestigious science journal Nature launched an informal survey into readers use of cognition-enhancing drugs, and found large-scale use (link). One in five respondents said they had used drugs for non-medical reasons to stimulate their focus, concentration or memory.

In 2008, Nature ran a commentary on this topic: Towards responsible use of cognitive enhancing drugs by the healthy. This article is well worth the time it takes to read. The authors outline the evidence in favor of the effectiveness of smart drugs and I will quote at length from the section Paths to Enhancement which reviews the nootropics known to boost brain power: Ritalin and Adderall

Many of the medications used to treat psychiatric and neurological conditions also improve the performance of the healthy. The drugs most commonly used for cognitive enhancement at present are stimulants, namely Ritalin (methyphenidate) and Adderall (mixed amphetamine salts), and are prescribed mainly for the treatment of attention deficit hyperactivity disorder (ADHD). Because of their effects on the catecholamine system, these drugs increase executive functions in patients and most healthy normal people, improving their abilities to focus their attention, manipulate information in working memory and flexibly control their responses Modafinil A newer drug, Modafinil (Provigil), has also shown enhancement potential. Modafinil is approved for the treatment of fatigue caused by narcolepsy, sleep apnoea and shift-work sleep disorder. It is currently prescribed off label for a wide range of neuropsychiatric and other medical conditions involving fatigue as well as for healthy people who need to stay alert and awake when sleep deprived, such as physicians on night call. In addition, laboratory studies have shown that modafinil enhances aspects of executive function in rested healthy adults, particularly inhibitory control. Unlike Adderall and Ritalin, however, Modafinil prescriptions are not common, and the drug is consequently rare on the college black market. But anecdotal evidence and a readers survey both suggest that adults sometimes obtain modafinil from their physicians or online for enhancement purposes. Aricept A modest degree of memory enhancement is possible with the ADHD medications just mentioned as well as with medications developed for the treatment of Alzheimers disease such as Aricept (donepezil), which raise levels of acetylcholine in the brain. Several other compounds with different pharmacological actions are in early clinical trials, having shown positive effects on memory in healthy research subjects. The authors focus at length on the potential risks and ethical concerns of using nootropic cognitive enhancers, but conclude: Like all new technologies, cognitive enhancement can be used well or poorly. We should welcome new methods of improving our brain function. In a world in which human workspans and lifespans are increasing, cognitive enhancement tools including the pharmacological will be increasingly useful for improved quality of life and extended work productivity, as well as to stave off normal and pathological age related cognitive declines23. Safe and effective cognitive enhancers will benefit both the individual and society. 3. Cortical Stimulation A number of studies in the last few years have shown very promising results from applying electrical current to the brain using a technology known as transcranial direct current stimulation (tDCS). tDCS is a noninvasive technique in which a weak current is applied to the brain constantly over time to excite or inhibit the activity of neurons.

In late 2010, a group of researchers from University College London and Oxford University published a study showing that tDCS applied to the parietal lobes enhanced a persons mathematical ability selectively, without influencing other cognitive functions. The improvement was found to have persisted six months after the training, showing the IQ gain was long-lasting. Earlier this year a study was published in Clinical Neurophysiology showing that tDCS of a the dorsolateral prefrontal cortex (dlPFC) improves working memory functioning. The dlPFC is a region in the frontal lobes toward the top and side: hence dorso (top) and lateral (side). The researchers report that there was significant improvement in speed of performance following tDCS on an n-back working memory task. In another study published earlier this year, a team at Centre for the Mind at the University of Sydney demonstrated that tDCS can dramatically improve insight problem solving. Three times as many cortically stimulated individuals succeeded in solving puzzles needing creative insight. People find it difficult to think outside of the box because their problem solving mind set becomes crystallized by past experience. By inhibiting the activity of the left temporal lobe, and stimulating activity in the right temporal lobe, this team changed the balance between the two hemispheres of the brain, leading to better release from mental sets and better creative insight. One of the team, Professor Snyder, believes brain boosting headgear could be widely used. The thinking cap of the future is not one that helps us to remember facts as the internet has solved that problem, but one that facilitates learning and unlearning mindsets. Its all about being original. Professor Snyder Some of the most recent work on tDCS was presented in September this year by Professor Prof Heidi Johansen-Berg and her colleagues at Oxford University. They found that just ten minutes of motor cortex brain stimulation increases the speed of learning motor skills. In their study a musical keyboard sequence was the learning task. While the stimulation didnt improve the participants best performance, the speed at which they reached their best was significantly increased. Professor Johansen-Berg

The researchers envisage the technique could be used to help in the training of athletes and suggest that the same method could be applied to other parts of the brain (such as the frontal or parietal cortex) to improve educational learning simply by positioning the electrodes in different locations so the current is focused on the correct area. The potential for self-experimentation is exciting. As this BBC report on cortical stimulation states: The relative simplicity, low price (around 2,000 per unit), and portability of the technology may mean that, following further research, a device could be designed to be automated for use at home. Research Summary One of my research areas is IQ and methods for increasing IQ. In this article I have reviewed three technologies that have been shown to have a substantial IQ increasing effect by the exacting standards of peer reviewed scientific research. The most effective technologies directly target working memory the general purpose RAM power of our brain. But technologies can be effectively applied in a targeted way to enhance more specialized aspects of cognitive function such as motor learning, numerical ability or insight problem solving. Intelligence augmentation is a cultural enterprise that is gaining momentum, but the technologies reviewed above take us into largely unexplored territory. The risks have not been fully quantified. It is our privilege to be in an era of both imaginative brain science, and biohackers responsible self-experimentation, to forge ahead in mapping out this territory in the spirit of pioneers. Please join me in this journey by subscribing to our blogs RSS feeds (right panel) or my CogPsyLab cognitive intervention research group for updates on our research and beta testing.