This instructable describes how to make a blue LED (470nm) transilluminator for DNA imaging using SYBR safe dyes. The transilluminator has a 6 x 7 cm viewing area for small agarose gels.
The transilluminator can also be modified to sit beneath the mini-gel electrophoresis tank (from a previous instructable), as described in Step 8. Used in this way, you can also visually track progress of your DNA during electrophoresis.
Open source hardware kit - This is an open source hardware project. To enable users to make or modify the device, I have included all of the PCB design files, enclosure design files and list of all hardware and electronics in a zip file attached below. We have also put together an LED Transilluminator Kit containing all of the parts described in this Instrucatble.
Step 1: Kit Contents
- Printed circuit board and electronics: DC jack, resistor, switch and 72 x blue LEDs
- Laser cut parts: 6 x black acrylic parts, 1 x clear acrylic parts, 1 x LED diffuser, 1 x blue light filter and an amber lid
- Enclosure hardware: 4 x male-female standoffs, 4 x 3/4" long standoffs, 4 x 1" long standoffs, 8 x screws and a mini-screwdriver
- Power supply. 12V power supply with an output current of at least 0.25A. Optional addition to the transilluminator kit. We use one from Jameco Cat # 252824.
Step 2: Soldering the Components Onto the LED Board
- 72 blue LEDs. Positions D1- D72. Make sure to check the orientation of the LEDs before soldering. The cathode of the LED is indicated by the flat edge on the silkscreen of the PCB. Insert the LED so that the short leg (cathode) is nearest the flat edge in the silk screen. See image.
- 5.6 Ohm resistor. Inset the resistor into either R1 or R2.
- DC jack. Insert into position P1.
- Switch. Insert into position SW1.
Step 3: Assembling the Enclosure Bottom
- Take the enclosure base and attach the four small male-female standoffs using four of the screws
- Place the PCB onto the 4 standoffs and screw in the 1" standoffs
- Take the four threaded studs and screw them all the way into the 1" standoffs
- Place the enclosure sides, enclosure back and enclosure front parts onto the enclosure base. Note that the four parts have short tabs on one side and longer tabs on the other side - place the parts so that the shorter tabs go into the base plate with the longer tabs are at the top
- The enclosure back part with the cutouts should fit over the DC jack and switch as shown in the images.
Step 4: Assembling the Enclosure Top
- First: Light diffuser film (2 sheets)
- Next: Blue filter
- Next: Black enclosure top (with the square cutout)
- Last: Clear enclosure top (with engraving)
Step 5: Assembling the Amber Top and Testing
- Screw the 4 remaining 3/4" standoffs onto the enclosure
- Place the amber lid on top and secure in place with the 4 remaining black screws.
Step 6: Preparing a SYBR Safe Agarose Gel
- Mini-gel electrophoresis tank or similar
- 1% agarose in TAE buffer (e.g. Melt-n-Pour Agarose, Carolina Biologicals, Cat # 21-7085)
- 1 x TAE buffer (e.g. diluted from 50x stock from Carolina Biologicals Cat # 21-9033)
- SYBR Safe DNA gel stain (Invitrogen Cat # 915548)
- DNA samples
- Seal the ends of the gel tray with the tape and place the gel comb in the slots of the gel tray. Place the tray on a flat surface.
- Melt the agarose in the microwave and let stand to cool until the bottle is warm enough to hold. Pour 50 mL of agarose into a small beaker.
- The SYBR safe stain is a 10,000 x stain. For 50 mL, add 5 µL of SYBR safe stain. Swirl the beaker to mix.
- Pour the agarose into the gel tray and let it cool and set (10 mins at room temp)
- Carefully remove the tape from the ends of the gel
- Place the tray in the tank filled with buffer
- Carefully remove the gel comb
- Pipet your DNA samples into the wells
- Place the lid onto the tank and connect the electrodes using banana cables
- Run the mini-gel at 50-100V. This takes approximately 30-60 mins.
Step 7: Viewing the Gel
Once the gel has finished running, take it out of the tank and place it onto the transilluminator. Switch on the LEDs - you should be able to see your DNA. In this image I have loaded 10-20 µL of a 1Kb DNA ladder (NEB QuickLoad, 50µg/mL).
Step 8: Viewing Gels During Electrophoresis
- Unscrew and remove the amber filter
- Unscrew and remove the 4 x threaded studs
- Place the 4 screws in each corner to hold the enclosure together
- Replace the clear tank lid with a lid cut from amber acrylic. Alternatively, place the amber from the transilluminator on top of the clear lid
- Place the transilluminator under the gel box. You should be able to see your DNA bands shortly after they have run into the gel.