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.
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.
led_transilluminator.zip374 KB
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Signing UpStep 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.
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congratulations to this instructable of yours. It is excellent, as the previous ones.
I have a question about DNA staining with SybrGreen. Apart from obvious health safety, are there any technical advantages or pitfalls, when compared to conventional Ethidium Bromide staining?
Isacco
That is a really good question. From what I know, the safety concern with ethidium bromide is really the big difference. In addition, UV can damage DNA which is not good obviously for cloning, sequencing etc. Having said that I used ethidium bromide for donkeys years in the lab and only recently switched to Sybr safe as I am no longer in an academic lab with access to endless piles of disposable gloves and convenient hazardous waste disposal.
As far as technical advantages and pitfalls, I should note that I use Sybr safe, not Sybr green. From what I have read from the literature, Sybr safe is reported to have a comparable sensitivity to ethidium bromide. I use the gel stain version - add it to the agarose before microwaving etc. so technically it handles very similar to ethidium bromide. I did notice that you can't re-use the gel though, which I have done with ethidium bromide gels. So far I haven't any problems with using Sybr safe. Cost is about $0.70 per gel versus pennies (?) for ethidium bromide. I would say it comes down to what you have access to in terms of waste disposal, type of transilluminator etc. and downstream application of your DNA where the advantages of not exposing your DNA to UV may be important.
I hope that helps !
I often see DNA with 254/302 nm UV transluminator and Etbr
I want to have one but I 'm in Vietnam
What materials of Blue filter ? Can i use blue arylic?
Thanks
I have another instructable on making a UV transilluminator for this purpose (http://www.instructables.com/id/UV-Transilluminator/) which works really well. However, as ethidium bromide and UV are not the friendliest tools to work with (especially in educational settings) I also wanted to make a blue light/sybr safe version.
For the blue filter, I tried a couple of different materials looking for something that can be laser cut and is not too expensive - this is blue acrylite # 668-0 which I bought online from acrylite-shop.com, part # 5C028GT.
I have just edited the enclosure bill of materials (found in the zip file in the Intro) to include a list of all of the acrylic with vendors/part numbers.
By the way, if you want a kit, I'm sure we can ship to Vietnam ;)
Many people will not appreciate the utility of this post but I'm so glad Instructables featured it in the newsletter. Our lab uses nucleic acid gel electrophoresis every single day so needless to say this peaked my interest.
This is a GREAT and thorough instructable, thank you for sharing! Great to include Sybr Safe dyes. I hope we can adopt this to replace our $20,000 "Gel-Doc" and out-dated Ethidium bromide dyes.
One addition I would make is an imaging system for record keeping. Have you seen the Berkeley made DIY Gel Doc? I built one up for our institute and it became pretty popular.
http://panic.berkeley.edu/~ghe/DIY_LowCostGelDoc/
Greg
Beth Israel Deaconess Med Ctr. Boston
Thanks for your comments ! Agree on the gel documentation step. I had seen the Berkeley DIY gel doc which is very nice, very cool that you built one. We are thinking about making an enclosure for this transilluminator with a camera mount and software for capturing and processing images. Do you use any image analysis software with your system such as ImageJ ?
We use the free Canon software called EOS Utility which includes Remote Shooting when the camera is connected to the computer by USB. You may be familiar with it as it was recommended in the Berkeley build.
EOS Utility comes free with Rebel cameras which is what we used for our GelDoc. Its getting harder to find (I think it doesnt come with PowerShot cameras) and not easy to download for free so it seems you usually have to get hold of a real disk.
2. Could you recommend a source for DNA ladders?
Good catch. I left off the electrophoresis power supply in the list of equipment !! I am still using the Whatman Biometra Model 125 low voltage power supply that I bought from Labrepco for $50. It has an output range of 50, 100 or 125 V. It is pictured on the mini-gel electrophoresis kit instructable. We are looking into an open source power supply for electrophoresis -- hopefully that will be the next Instructable !
I can recommend the DNA ladders from NEB (www.neb.com). I have also used the ladders from Promega but recently switched to the NEB QuickLoad 1 Kb DNA ladder for convenience and cost. I recently ordered catalog # N0468S, which with shipping came to $82. Not very cheap, so I use it sparingly :)
Good luck.
No, it's not obvious to us and No, I'm not picking on this in particular; most instructable's are not discernable as to what they do or what purpose this thing is used for which ultimately would decide whether we build one or not.
This would also be a great teaching tool for high school biology. Kudos to the author.
Although, I am not singling this instructable out, its a good instructable, I just want to see it used in a practical application in order to understand what exactly it can be used for.
Many instructables focus too heavily on the building of the subject because they know its use and the limitation or the vast possibilities of it, but if you don't know what those limitations are, or indeed the practical use or even the potential practical use of the instructable the audience who do not know how it can be used is left if its worth doing.