What is Loomino?
Loomino is a simple DNA synthesizer that is built using open-source software and hardware. Its mission is to reduce the amount of time a lab would need to wait for synthesized DNA to arrive. Current DNA synthesizers use phosphoramidite-based procedures with hazardous chemicals and dangerous waste. It is highly uncommon that a lab outside of large corporations would be able to perform DNA synthesis, and therefore this process slows down bioresearch.
The machine itself has 3 subsystems: fluidics, thermal cycling, and electronics. The main criteria we were aiming for was ease of use, and feasibility. This criteria was used to judge the design, and optimize the machine.
Fluidics
In order to properly add reagents to the reaction chamber, Loomino uses a series of peristaltic pumps to apply pressure thus creating flow. These peristaltic pumps are constructed out of a NEMA17 stepper motor, 3D printed parts, and machined bearings. As peristaltic pumps are expensive, we decided that an open source design would be best suited for Loomino. The overall cost of such a pump is far cheaper than purchasing a prefabricated pump from a manufacturer.
Another advantage of using a peristaltic pump is that Loomino has the ability to draw air into the tubes. This allows for a gas buffer between the fluids which means we are able to move accurate volumes of fluid without excess waste. A mechanism is able to lower the reagents such that the tubes are not submerged in fluid, allowing for us to draw air. Without such a mechanism, large amounts of reagent would be wasted during the wash process which reduces the cost efficiency of the device.
There were many alternatives to create fluid flow such as using a system of syringes, or having a gas pressure microfluidic device. The advantage of having a syringe system is that the mechanism needed to pump is relatively simple. By rotating a threaded rod with a motor, we could create linear motion that would compress a syringe, thereby pumping fluids. The issue is that it lacks the ability to pump air, and thus we would encounter large amounts of waste. Also, a primary concern is accuracy. As the stepper motors are able to move within a 1.8 degree rotation excluding half-steps, we are able to more finely control flow rates. A microfluidic device requires more equipment either for lithography or molding PDMS. A gas pressure system would also need to be implemented, and therefore we determined these to be not feasible.
Image of the peristaltic pump.
Thermal Cycling
De Novo Synthesis requires the removal of a heat-labile group at 95 degrees C. The reaction chamber must then be cooled in preparation for the next reagent for the heating process. This means Loomino must cycle through temperatures of 95 degrees C and 37 degrees C with relative accuracy and speed. In order to accomplish the heating process, Loomino is equipped with a heating cartridge that quickly raises the temperature of the reaction chamber. A thermistor is attached to regulate the temperature and provide feedback for the system.
Once the reaction has completed in the heating step, the cooling process begins. A thermoelectric cooler is turned on, rapidly cooling the reaction chamber. A stock fan and heat sink are attached as well to assist in cooling and providing airflow through the machine. Once the machine drops to the proper temperature, the cooling stops. The thermistor continues to provide feedback and regulate the temperature.
Once De Novo Synthesis has completed, the machine is able to run the same thermal cycling process as a PCR machine. Loomino is able to cycle between 95 degrees C, 55 degrees C, and 72 degrees C for the standard PCR cycle. As it is electronically controlled, any number of cycles is possible.
Heating cartridge used in the heating element.
Electronics
Loomino is controlled using an Arduino microcontroller. It is equipped with motorshields to send the proper signals to control the stepper motors. The system is preprogrammed to initiate the proper procedure to run de novo synthesis. The only constant variation control is in the thermal cycling. The thermistor uses variable resistance based on temperature to change the current running through the circuit. That current is then translated to a temperature. Using proportional-integral-derivative control, the Arduino is able to turn the heating cartridge on and off to remain at a stable temperature.
Bill of Materials
| FLUIDICS - PERISTALTIC PUMP | CASUN STEPPING MOTOR | 42SHD0411-20B | NEMA17 STEPPER MOTOR, 42mm, 4.4 kg/cm | 7 | Higher torques above 4.4 kg/cm are acceptable |
| FLUIDICS - PERISTALTIC PUMP | 3D PRINTED BEARING HUB | LMNO-PP-3DBH | BEARING HUB holds bearings in place for peristaltic pump rotations | 7 | Flexibility will determine how well the tubing is squeezed. Leads to pressure differences. |
| FLUIDICS - PERISTALTIC PUMP | 3D PRINTED PUMP CASING | LMNO-PP-3DPC | PUMP CASING is the outer rim that encases the pump and provides a wall to squeeze the tube | 7 | |
| FLUIDICS - PERISTALTIC PUMP | 3D PRINTED TUBE CLAMP | LMNO-PP-3DTC | TUBE CLAMP keeps tubing from slipping from peristaltic pump | 14 | |
| FLUIDICS - PERISTALTIC PUMP | BRASS BEARING | F693ZZ | 3mm x 8mm x 4 mm BRASS BEARING used to apply pressure to tube. Attached to bearing hub | 28 | |
| FLUIDICS - PERISTALTIC PUMP | M3 SCREW | 91290A111 | Black-Oxide Class 12.9 Socket Head Cap Screw, Alloy Steel, M3 Thread, 6mm Length, 0.50mm Pitch | 35 | |
| FLUIDICS - PERISTALTIC PUMP | M4 SCREW | 91290A168 | Black-Oxide Class 12.9 Socket Head Cap Screw, Alloy Steel, M4 Thread, 20mm Length, 0.70mm Pitch | 14 | Similar sizes may be used. Sizes may be adjusted. |
| FLUIDICS - PERISTALTIC PUMP | M3 HEX NUT | 90695A033 | Class 04 Steel Thin Hex Nut - DIN 439B, Zinc Plated, M3x0.5 Thread Size, 5.5mm Wide, 1.8mm High | 7 | |
| THERMAL CYCLER | THERMOELECTRIC COOLER | TEC1-12706 | 40mm x 40mm x 3.6mm 12V 92W Peltier Cooler | 1 | Various peltier devices may be used |
| THERMAL CYCLER | HEATSINK | AMD Socket AM2 Aluminum Heat Sink | 1 | Similar heatsinks can be used. Edits to CAD files must be made to accomodate. | |
| THERMAL CYCLER | CPU FAN | AMD Socket AM2 2.75" Fan | 1 | Match CPU FAN to heatsink | |
| THERMAL CYCLER | 3D PRINTED PELTIER CASE | LMNO-TC-3DPC | 3D PRINTED CASE to hold thermoelectric cooler | 1 | Match PELTIER CASE to heatsink |
| THERMAL CYCLER | 3D PRINTED OVERHEAD CASE | LMNO-TC-3DOC | 3D PRINTED CASE to hold heating element | 1 | |
| THERMAL CYCLER - HEATING ELEMENT | HEATING CARTRIDGE | 0710270192984 | REPRAP 12V 40W Heating Cartridge used for 3D printers | 1 | |
| THERMAL CYCLER - HEATING ELEMENT | THERMISTOR | 0087169815454 | M3 Stud THERMISTOR for Reprap 3D printer extruder | 1 | |
| THERMAL CYCLER - HEATING ELEMENT | ALUMINUM HEAT BLOCK | LMNO-TC-MAHE | 20mm x 40mm x 10mm ALUMINUM HEAT BLOCK to transmit heat from heating cartridge. Machined. | 1 | |
| THERMAL CYCLER | M8 SCREW | 90854A180 | High-Strength Zinc Yellow-Chromate Plated Cap Screw, Class 10.9 Steel, M8 Full Thread, Pitch: 1.25, 60mm Long | 4 | |
| THERMAL CYCLER | M8 HEX NUT | A000066 | ARDUINO UNO REV3 ATmega328 microcontroller | 1 | |
| ELECTRONICS | ARDUINO UNO REV3 | 90695A040 | Class 04 Steel Thin Hex Nut - DIN 439B, Zinc Plated, M8x1.25 Thread Size, 13mm Wide, 4mm High | 4 | |
| ELECTRONICS | MOTORSHIELD v2.3 | Adafruit Motor/Stepper/Servo Shield for Arduino v2 Kit - v2.3 | 2 | ||
| FLUIDICS | TUBING | 5054K64 | Metric High-Temperature Silicone Rubber Tubing, Soft, OD 3mm, ID 1mm | 5 ft |
CAD Files can be found here
By clicking this link, you will download a .zip file with the compressed Solidworks files.