Difference between revisions of "Team:UMaryland/Design"

Revision as of 22:04, 18 September 2015

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UMD DIY PCR

How we created a dual purpose PCR machine and incubator out of a hair dryer.

Background

The PCR machine is a common machine used in biological laboratories to amplify or extend fragments of DNA to be used in subsequent experiments. This tool is especially relevant to iGEM and SynBio labs who pave the way to vaster applications of We began this project with the vision to create a machine that would be

Our first design for a DIY PCR machine was modeled after a more conventional PCR machine. This first prototype relied on two Peltier units stacked on top of each other to heat a customized aluminum block that held the PCR tubes. In order for the system to have feedback, we embedded a temperature sensor in the aluminum block to measure the temperature of the PCR tube wells. The sensor then reported back to an Arduino UNO, which then regulated the energy flow to the Peltier units, thereby regulating the temperature of the block and tubes. However, after much testing, this design proved to be unoriginal, expensive, and inefficient. While the conventionality of the design itself did not pose an issue, we realized that the parts used to assemble it were not as well-known or easily accessible to the general public, which we felt would take away from the possible applications of this machine. In addition, although the price of this first prototype was relatively inexpensive in contrast to laboratory grade PCR machines, the price still ranged in the hundreds of dollars. Finally, the greatest issue with our design was the inefficiency of the hardware; we found that the Peltier units were not able to quickly cycle through the desired temperatures, causing the unit to take 5 to 10 minutes just to rise up to 95℃. After considering all of these factors, we began a redesign of our machine to better suit the needs of the DIY market.

The idea for our current thermocycler design first came into form when we found that our original prototype was not ramping up to the desired temperatures fast enough. We thus looked into other options such as the heating element in a hair dryer. We found that the hair dryer was able to reach very high temperatures—much higher than the desired maximum of 95℃ for PCR—in a matter of seconds. We then made a decision to suspend construction on the Peltier-centered thermocycler in order to see how successful we could be with making a rapid PCR machine out of a hair dryer. Before this decision, we took into consideration the danger of working with a hair dryer, failure due to uncertainty that the machine could be effectively controlled, and, on top of that, having less time to work on it. Nevertheless, we took the risk.
Please continue on to see the design of our machine.

What is PCR?

Polymerase Chain Reaction or PCR is a common tool used in the field of biology to amplify DNA or RNA. Invented by Dr. Kary Mullis, PCR is conducted trough cycling DNA, primers and polymerase through various temperatures. The reaction is started by heating the reaction mix to 95 degrees Celsius. The high heat overcomes base stacking interactions and hydrogen bonds which maintain the double helix, a process called denaturation. The machine then cools down to an annealing temperature in order for primers, short ssDNA oligos, to recognize selected DNA sequences, form duplex, and allow for polymerase to bind. Annealing is followed by extension, which is performed by the polymerase at its active temperature, typically around 72 degrees. The polymerase forms a daughter strand by adding nucleotides to the primer in the 5'-3' direction. I don't think this is necessary, especially not here. If you want to write how PCR works, put it in description. PCR is also a very complicated process so we'll need to invest a lot of space into it if you want to do it justice

Construction Hazard. Build at your own risk.

UMD DIY PCR

Our first design for a DIY PCR machine was modeled after a more conventional PCR machine design. This first prototype consisted of two Peltier units stacked on top of each other that would then heat a customized aluminum block that sat on top of the two units and held the PCR tubes. In order for the system to have feedback, we embedded a temperature sensor in the aluminum block to measure the temperature of the PCR tube wells. The sensor then reported back to an Arduino UNO, which then regulated the energy flow to the Peltier units, thereby regulating the temperature of the block and tubes. However, after much testing, this design proved to be unoriginal, expensive, and inefficient. While the conventionality of the design itself did not pose an issue, we realized that the parts used to assemble it were not as well-known or easily accessible to the general public, which we felt would take away from the possible applications of this machine. In addition, although the price of this first prototype was relatively inexpensive in contrast to laboratory grade PCR machines, the price still ranged in the hundreds of dollars. Finally, the greatest issue with our design was the inefficiency of the hardware; we found that the Peltier units were not able to quickly cycle through the desired temperatures, causing the unit to take 5 to 10 minutes just to rise up to 95℃. After considering all of these factors, we began a redesign of our machine to better suit the needs of the DIY market.

@@ Line 375: / Line 375: @@
   &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#000000">laststep</font><font color="#434f54">++</font><font color="#000000">;</font>
   &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#000000">}</font>
-  &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#434f54">//If the pcr has maintained the setpoint for the desired amount of time, it will enter this loop and begin cooling</font>
+  &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#7e7e7e">/*If the pcr has maintained the setpoint for the desired amount </font><font color="#7e7e7e"> &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;of time, it will enter this loop and begin cooling*/</font>
   &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#5e6d03">if</font> <font color="#000000">(</font><font color="#000000">(</font><font color="#d35400">millis</font><font color="#000000">(</font><font color="#000000">)</font> <font color="#434f54">&gt;</font> <font color="#000000">(</font><font color="#000000">t1Melt</font> <font color="#434f54">+</font> <font color="#000000">cycleStart</font><font color="#000000">)</font><font color="#000000">)</font> <font color="#434f54">&&</font> <font color="#000000">(</font><font color="#000000">laststep</font> <font color="#434f54">==</font> <font color="#000000">stepnow</font><font color="#000000">)</font><font color="#000000">)</font> <font color="#000000">{</font>
   &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#000000">stepnow</font><font color="#434f54">++</font><font color="#000000">;</font>
@@ Line 394: / Line 394: @@
   &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#000000">laststep</font><font color="#434f54">++</font><font color="#000000">;</font>
   &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#000000">}</font>
-  &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#434f54">//checks if the machine has maintained the temperature for the desired amount of time, and begins the next step</font>
+  &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#434f54">//checks if the machine has maintained the temperature for </font>
+ &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#434f54">//the desired amount of time, and begins the next step</font>
   &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#5e6d03">if</font> <font color="#000000">(</font><font color="#000000">(</font><font color="#d35400">millis</font><font color="#000000">(</font><font color="#000000">)</font> <font color="#434f54">&gt;</font> <font color="#000000">(</font><font color="#000000">t1PCR</font> <font color="#434f54">+</font> <font color="#000000">cycleStart</font><font color="#000000">)</font><font color="#000000">)</font> <font color="#434f54">&&</font> <font color="#000000">(</font><font color="#000000">laststep</font> <font color="#434f54">==</font> <font color="#000000">stepnow</font><font color="#000000">)</font> <font color="#434f54">&&</font> <font color="#000000">(</font><font color="#000000">cycle</font> <font color="#434f54">==</font> <font color="#000000">2</font><font color="#000000">)</font><font color="#000000">)</font> <font color="#000000">{</font>
   &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#000000">stepnow</font><font color="#434f54">++</font><font color="#000000">;</font>
@@ Line 412: / Line 413: @@
   &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#000000">laststep</font><font color="#434f54">++</font><font color="#000000">;</font>
   &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#000000">}</font>
-  &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#434f54">//checks if the machine has maintained the temperature for the desired amount of time, and begins the next step</font>
+  &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#7e7e7e">/*checks if the machine has maintained the temperature </font><font color="#7e7e7e"> &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;* for the desired amount of time, and begins the next step*/</font>
   &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#5e6d03">if</font> <font color="#000000">(</font><font color="#000000">(</font><font color="#d35400">millis</font><font color="#000000">(</font><font color="#000000">)</font> <font color="#434f54">&gt;</font> <font color="#000000">(</font><font color="#000000">textension1</font> <font color="#434f54">+</font> <font color="#000000">cycleStart</font><font color="#000000">)</font><font color="#000000">)</font> <font color="#434f54">&&</font> <font color="#000000">(</font><font color="#000000">laststep</font> <font color="#434f54">==</font> <font color="#000000">stepnow</font><font color="#000000">)</font><font color="#000000">)</font> <font color="#000000">{</font>
   &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#000000">pcr</font> <font color="#434f54">=</font> <font color="#00979c">"heating to melt "</font><font color="#000000">;</font>
@@ Line 466: / Line 467: @@
   &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#000000">}</font>
   &nbsp;&nbsp;&nbsp;<font color="#000000">}</font>
-  &nbsp;<font color="#7e7e7e">/*</font><font color="#7e7e7e"> &nbsp;&nbsp;* Begins the last step, which again works in a similar fashion to the other steps, but may differ in the </font><font color="#7e7e7e"> &nbsp;&nbsp;* final extension temperature</font><font color="#7e7e7e"> &nbsp;&nbsp;*/</font>
+  &nbsp;<font color="#7e7e7e">/*</font><font color="#7e7e7e"> &nbsp;&nbsp;* Begins the last step, which again works in a similar fashion </font><font color="#7e7e7e"> &nbsp;&nbsp;* to the other steps, but may differ in the final extension temperature</font><font color="#7e7e7e"> &nbsp;&nbsp;*/</font>
   &nbsp;<font color="#000000">}</font> <font color="#5e6d03">else</font> <font color="#5e6d03">if</font> <font color="#000000">(</font><font color="#000000">(</font><font color="#000000">stepnow</font> <font color="#434f54">&gt;</font><font color="#434f54">=</font> <font color="#000000">cyclenum</font> <font color="#434f54">&&</font> <font color="#000000">stepnow</font> <font color="#434f54">&lt;</font><font color="#434f54">=</font> <font color="#000000">cyclenum</font> <font color="#434f54">+</font> <font color="#000000">2</font> <font color="#000000">)</font><font color="#000000">)</font> <font color="#000000">{</font>
   &nbsp;&nbsp;&nbsp;<font color="#5e6d03">if</font> <font color="#000000">(</font><font color="#000000">(</font><font color="#000000">(</font><font color="#000000">stepnow</font> <font color="#434f54">%</font> <font color="#000000">3</font><font color="#000000">)</font> <font color="#434f54">==</font> <font color="#000000">1</font><font color="#000000">)</font><font color="#000000">)</font> <font color="#000000">{</font>
@@ Line 513: / Line 514: @@
   &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<font color="#000000">}</font>
   &nbsp;&nbsp;&nbsp;<font color="#000000">}</font>
-  &nbsp;<font color="#7e7e7e">/**</font><font color="#7e7e7e"> &nbsp;&nbsp;* Once the last step has been completed, determined by stepnow being greater than cyclenum+2,</font><font color="#7e7e7e"> &nbsp;&nbsp;* the code will tell the pcr to hold at 4 C. It will continue looping at this step indefinitely</font><font color="#7e7e7e"> &nbsp;&nbsp;* because the conditions will no longer satisfy any of the other if statement</font><font color="#7e7e7e"> &nbsp;&nbsp;*/</font>
+  &nbsp;<font color="#7e7e7e">/**</font><font color="#7e7e7e"> &nbsp;&nbsp;* Once the last step has been completed, determined by </font><font color="#7e7e7e"> &nbsp;&nbsp;* stepnow being greater than cyclenum+2,the code will tell the </font><font color="#7e7e7e"> &nbsp;&nbsp;* pcr to hold at 4 C. It will continue looping at this step indefinitely</font><font color="#7e7e7e"> &nbsp;&nbsp;* because the conditions will no longer satisfy any of the other if statement</font><font color="#7e7e7e"> &nbsp;&nbsp;*/</font>
   &nbsp;<font color="#000000">}</font> <font color="#5e6d03">else</font> <font color="#000000">{</font>
   &nbsp;&nbsp;&nbsp;<font color="#00979c">int</font> <font color="#000000">strt</font> <font color="#434f54">=</font> <font color="#000000">1</font><font color="#000000">;</font>
@@ Line 528: / Line 529: @@
   &nbsp;&nbsp;&nbsp;<font color="#000000">}</font>
-  &nbsp;&nbsp;&nbsp;<font color="#434f54">//Once the pcr has reached 37 C, the pcr will print out that the pcr has shutt off</font>
+  &nbsp;&nbsp;&nbsp;<font color="#7e7e7e">/*Once the pcr has reached 37 C, the pcr will </font><font color="#7e7e7e"> &nbsp;&nbsp;&nbsp;print out that the pcr has shutt off*/</font>
   &nbsp;&nbsp;&nbsp;<font color="#5e6d03">if</font> <font color="#000000">(</font><font color="#000000">(</font><font color="#000000">Input</font> <font color="#434f54">-</font> <font color="#000000">38</font> <font color="#434f54">+</font> <font color="#000000">1</font><font color="#000000">)</font> <font color="#434f54">&lt;</font> <font color="#000000">0</font><font color="#000000">)</font> <font color="#000000">{</font>
   &nbsp;&nbsp;&nbsp;&nbsp;<font color="#d35400">digitalWrite</font><font color="#000000">(</font><font color="#000000">signalr1</font><font color="#434f54">,</font> <font color="#00979c">HIGH</font><font color="#000000">)</font><font color="#000000">;</font>
@@ Line 551: / Line 552: @@
   &nbsp;&nbsp;&nbsp;<font color="#d35400">digitalWrite</font><font color="#000000">(</font><font color="#000000">signalr2</font><font color="#434f54">,</font> <font color="#00979c">LOW</font><font color="#000000">)</font><font color="#000000">;</font>
   &nbsp;&nbsp;&nbsp;<b><font color="#d35400">Serial</font></b><font color="#434f54">.</font><font color="#d35400">println</font><font color="#000000">(</font><font color="#00979c">"COOLING"</font><font color="#000000">)</font><font color="#000000">;</font>
-  <font color="#434f54">// if the pcr machine's temperature reaches 97, cooling will automatically be initiated and a death message will be displayed</font>
+  <font color="#7e7e7e">/* if the pcr machine's temperature reaches 97, cooling will </font><font color="#7e7e7e"> &nbsp;&nbsp;&nbsp;automatically be initiated and a death message will be displayed*/</font>
   &nbsp;<font color="#000000">}</font><font color="#5e6d03">else</font> <font color="#5e6d03">if</font> <font color="#000000">(</font><font color="#000000">(</font><font color="#000000">Input</font> <font color="#434f54">-</font> <font color="#000000">deathT</font> <font color="#434f54">+</font><font color="#000000">1</font><font color="#000000">)</font> <font color="#434f54">&gt;</font> <font color="#000000">0</font><font color="#000000">)</font><font color="#000000">{</font>
   &nbsp;&nbsp;&nbsp;<font color="#d35400">digitalWrite</font><font color="#000000">(</font><font color="#000000">signalr1</font><font color="#434f54">,</font> <font color="#00979c">HIGH</font><font color="#000000">)</font><font color="#000000">;</font>
@@ Line 557: / Line 558: @@
   &nbsp;&nbsp;&nbsp;<b><font color="#d35400">Serial</font></b><font color="#434f54">.</font><font color="#d35400">println</font><font color="#000000">(</font><font color="#00979c">"DEATH"</font><font color="#000000">)</font><font color="#000000">;</font>
-<font color="#7e7e7e">/* these two statements work on the maintaining the temperature, by controlling heating and cooling when there is overshoot or undershoot </font><font color="#7e7e7e"> &nbsp;&nbsp;in the temperature*/</font>
+<font color="#7e7e7e">/* these two statements work on the maintaining the temperature, by </font><font color="#7e7e7e"> * controlling heating and cooling when there is overshoot or undershoot </font><font color="#7e7e7e"> &nbsp;&nbsp;in the temperature*/</font>
   &nbsp;<font color="#000000">}</font> <font color="#5e6d03">else</font> <font color="#5e6d03">if</font> <font color="#000000">(</font><font color="#000000">Output</font> <font color="#434f54">&gt;</font> <font color="#000000">now</font> <font color="#434f54">-</font> <font color="#000000">windowStartTime</font> <font color="#434f54">-</font> <font color="#000000">1</font><font color="#000000">)</font> <font color="#000000">{</font>
   &nbsp;&nbsp;&nbsp;<font color="#d35400">digitalWrite</font><font color="#000000">(</font><font color="#000000">signalr1</font><font color="#434f54">,</font> <font color="#00979c">HIGH</font><font color="#000000">)</font><font color="#000000">;</font>

Difference between revisions of "Team:UMaryland/Design"

Revision as of 22:04, 18 September 2015

Warning banner up here

UMD DIY PCR

Background

What is PCR?

UMD DIY PCR

Design

Hardware

Problems and Current issues

CODE