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
Each blue LED transilluminator kit comes with the following parts: 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. Note: A complete list of all of the parts for the enclosure and PCB are included in the zip file in the previous step.
Step 2: Soldering the components onto the LED board
Solder the following through-hole components onto the LED Transilluminator PCB. Note: for an Instructable on soldering components onto circuit boards, check out How-to-solder (Steps 4& 5) by noahw. 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. Once all the components are in place, plug in the 12 V power supply and switch on the board to check all of the LEDs are working correctly.
Step 3: Assembling the enclosure bottom
Before starting assembly, peel off any of the paper backing from the laser cut parts.
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
Place the next 4 laser cut parts in the following order: First: Light diffuser film (2 sheets) Next: Blue filter Next: Black enclosure top (with the square cutout) Last: Clear enclosure top (with engraving) Edit: A list of all of the acrylic materials used in the enclosure and the filters are listed in the file "BOM_enclosure.txt" in the "enclosure" folder of the zip file (see Intro). List includes suggested vendors and part numbers.
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. You have finished assembly of your transilluminator ! To test the transilluminator, plug in the power supply and switch it on.
Step 6: Preparing a SYBR safe agarose gel
Materials and Equipment 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) Tape Micropipette DNA samples Methods
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
The transilluminator can be easily modified to use used with the mini-gel electrophoresis tank to view DNA 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.