New Home BioLab Advice

[Bryan Daniels]

Many, many years ago, I was a chemistry major, but never pursued the sciences professionally. Recently, I have been availing myself of some amazing online courses, including excellent biology courses from MIT on Edx. (I just finished a great class on DNA Replication and Repair and looking forward to one on Transcription and Transposition.) Those classes have inspired me to start my own home lab for me and my kids. I have the basics: miniPCR, electrophoresis chamber, transilluminator, centrifuge, pipettes, scales, heat, cold, glassware etc. I have been focused on “Hello World” applications: Forensic Kits, GMO testing of foods, bar-coding, etc.

I am now interested in cranking it up a level. I have been reading through Molecular Biology Techniques (http://www.amazon.com/Molecular-Biology-Techniques-Third-Laboratory/dp/0123855446/ref=sr_1_1?ie=UTF8&qid=1445102583&sr=8-1&keywords=molecular+biology+techniques) and would now like to attempt some of the experiments and assays described in that book. (I have been emailing Josiah Zayner, who has begun helping me out. Thanks Josiah!)

I've been reading a fair amount about the need to quantify DNA for various assays. I keep hearing about 260/280 UV Spectroscopy suggesting the Nanodrop (and recently the DeNovix DS-11). Sadly, even used, the Nanodrop is a $5,000 piece of equipment. How do diybio labs quantify DNA? If so, what are the DIYbio solutions?

I have a miniPCR, which works great for me. Related to the first question, I've been reading about the new openqPCR device. Any thoughts or experiences with it? I may be mixing apples and oranges, but might it be used not only as a qPCR, but also to quantify DNA? I guess it probably still wouldn't be able to validate purity.

I would like to get an autoclave. Once again, I am looking for a reasonable solution. I've seen the nice laboratory autoclaves on the one hand and the pressure-cooker "crock pots" on the other. Not sure how well the later perform. Is this an area to spend the extra dollars?

I am excited to be a part of the diybio movement and have read quite a few posts posted in this community.

Thanks very much in advance for any help or ideas.

Bryan Daniels

[Bryan Jones]

It's more tedious than a uv/vis reading, but you can quantify a homogeneous DNA sample with an agarose gel by including standards of known concentration. Probably won't help much with complex samples like a genomic DNA prep though.

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[Koeng]

I use a pressure cooker for an autoclave. 

DNA quantification is usually just done as Bryan Jones said, on an agarose gel. http://www.geneandcell.com/collections/open-source is an example of some ladders, though admittably I'm still working through the ladder NEB sends with their enzymes. What is your purpose in quantifying DNA? If you're cloning, pick up some HiFi mastermix (or make the gibson mastermix yourself). Just purify the DNA after a PCR reaction ( http://nebuilder.neb.com/ for primers) and you're set. 

-Koeng

[Sebastian S Cocioba]

If you want to just assemble DNA, try it the old fashioned way and use cheap restriction enzymes and an open source vector like p2KB. Traditional cloning is sooo much cheaper and the only catch is time...which most DIYBio labs have in most abundance. I use NEB's Cutsmart enzymes and just miniprep, cut 16uL of plasmid with no quantification whatsoever and I get my clone 8/10 times. I bought a Qubit 2.0 which is an awesome doodad but I find its use for cloning a single part at a time to be overkill. Goldengate and gibson are great but $10/rxn is a bit steep in the long run. I just take my time and get a couple guaranteed clones per week with plenty of time to plan and think in between runs. For certain cases the old style is best when starting up. I can forward you my protocols for cutting and pasting and cell prep if you would like. Glad to hear you are getting your kids involved too! Best of luck!

Sebastian S. Cocioba
CEO & Founder
New York Botanics, LLC