Take the 2-minute tour ×
Biology Stack Exchange is a question and answer site for biology researchers, academics, and students. It's 100% free, no registration required.

There's undoubtedly more than one way to do this, but if a DIY biologist were to attempt to detect food fraud (e.g. as done by students from Stanford University and Trinity School, Manhattan with respect to fish samples from markets and sushi restaurants), then what would be the minimum steps and equipment?

(I know barely anything about molecular genetics, but have been reading about DremelFuge, OpenPCR, and Blue Transilluminator, and wondered whether they - or things like them - might get such an investigator some of the way towards the goal above; and what else would be required.)

share|improve this question
    
In principle this can be done at home. How familiar are you with PCR? –  Chris May 7 at 7:19
    
@Chris, I understand in outline what PCR is supposed to do, but not in detail; nor have I ever performed PCR myself. –  sampablokuper May 7 at 13:20
    
I have described the general process below, but I don't think detailed protocols belong on this site. (But perhaps it would be OK to write them in tag-wikis?). Even with very detailed protocols, there is often quite a bit of expected knowledge that is not mentioned, though. The best solution is perhaps to get some hands-on training with someone knowledgable. That said, neither PCR nor gel electrophoresis is particularly difficult, so you might be able to do it without instruction, but expect quite a bit of blundering in the start! –  jarlemag May 7 at 16:45
add comment

2 Answers 2

up vote 6 down vote accepted

There are several ways you could go about identifying species through DNA. If you want to do everything yourself, the simplest option in terms of equipment needed consists of evaluating fragment lengths observed during gel electrophoresis after amplifying specific DNA sequences using PCR.

If you are content with some outsourcing, you can also send DNA samples to a commercial company for sequence analysis.

A compromise between these options in terms of information obtained, is to do study Restriction Fragment Length Polymorphism (RFLP) by amplifying DNA fragments and using restriction enzymes to cut the fragments, before analyzing the fragmentation pattern using gel electrophoresis. To perform RFLP analysis, you would need to obtain restriction enzymes in addition to the chemicals mentioned below, and they can be a bit pricey.

The minimum equipment would consist of a PCR machine, one or more pipettes with matching pipette tips, a gel electrophoresis tray with power supply and a transilluminator (preferably blue-light/non-UV). A centrifuge is not strictly necessary, but can be useful for processing/filtering your DNA source.

Some chemicals will also be needed: Polymerase and dNTPs for the PCR reaction (or a pre-made "master mix" containing both), electrophoresis-grade agarose and running buffer for the electrophoresis, along with a DNA dye specific to the type of transilluminator (Usually UV or blue light). For a blue-light transilluminator, GelGreen is a suitable DNA dye. You will also want to use a "loading dye" to mix in your DNA sample before applying it to the electrophoresis gel. This can either be purchased or prepared yourself by mixing sugar and food coloring in water.

You need some form of heating to dissolve the agarose - a microwave oven is convenient for this, but take care to avoid over-heating, glass explosions or flash boiling. It is convenient but not strictly necessary to have some lab glassware. Preferably use a screw-top bottle to mix your agarose solution. Always leave the top off when heating bottles.

Photographic equipment can be also useful for documenting results of gel electrophoresis.

Finally, you will need single-stranded DNA oligomers (primers) specific to the DNA regions you want to amplify. DNA primers can be bought from a number of companies, but it varies how easy it is for non-affiliated individuals to order and make payments. Macrogen has been my choice: They both deliver DNA primers and perform DNA sequencing.

You may be interested in the following thread on the DIY Bio e-mail group: https://groups.google.com/forum/#!topic/diybio/cPzfEuiZH58

I have collected some of the primer sequences mentioned in the thread on a page on OpenWetware: http://openwetware.org/wiki/User:Jarle_Pahr/Meat

share|improve this answer
    
Thanks :) Maybe I just haven't read enough, but is there a DIY protocol that would allow the detection of material from a completely unexpected species? My current understanding is that one needs a PCR primer (or perhaps restriction enzyme) appropriate to each species one wishes to test for: e.g. the only way to detect, say, tuna fish in dodgy "beef" bolognese would be to use a primer suitable for amplifying tuna DNA fragments, but this would not detect haddock present in the same sample. Is there a DIY protocol that would detect the haddock without specifically testing for it? –  sampablokuper May 9 at 3:03
add comment

This can in principle be done at least partly with DIY methods, by using PCR followed by gel electrophoresis. The DremelFuge, OpenPCR, and Blue Transilluminator would be the primary tools, along with pipettes, test tubes, autoclave (or equivalent), etc.

For a clear video demonstration of using PCR to amplify DNA from various samples, as well as a PDF containing the details of the protocol used, see www.dnabarcoding101.org. That procedure would be followed by using gel electrophoresis on a small portion of the amplified product, to check that the product appears to be of adequate quality for sequencing. If so, then the remaining portion of the product can be sent to a lab for sequencing.

Alternatively, by using a suitable primer for each species one wishes to test for, one could check for the presence of that species using PCR and gel electrophoresis alone.

share|improve this answer
add comment

Your Answer

 
discard

By posting your answer, you agree to the privacy policy and terms of service.

Not the answer you're looking for? Browse other questions tagged or ask your own question.