DIYBIOFTW

Gelfoo v4 - ScixKsr workshop design

Hi Everybody!

I'm Mac. I'm a biotech maker. I help run diybio.org and have a modest online retail site where I sell affordable pipettes for molecular biology. My goal is to make the equipment toolchain for doing basic molecular biology more accessible through radically affordable prices and better industrial design.

Genefoo gelfoo concept.

A web-enabled system for gel-documentation. Posts images to dropbox, evernote. Hypothesis: users waste time getting images from camera to lab notebook.

Gels?

Gel electrophoresis is a routine (i.e. daily) technique used in molecular biology labs to separate DNA molecules from one another based on differences in their length. A 1.5% agarose gel can discriminate DNA fragments with lengths between 200 base pairs (bp) and 3000 bp (3 kb). To run and image a gel, the following equipment is needed:

agarose, dh2o, gel buffer (TBE, TAE), DNA binding dye, running & loading buffer

gel casting tray, submarine gel box, comb

power supply - establishes 100-300v electric field across gel

transilluminator - excites bound DNA dye

"geldoc" imager - box that fits on top of transilluminator & holds a camera with filter. The filter cuts out the transilluminator light and passes the light from the fluorescing DNA-bound dye.

Several open-source kits and designs exist for most of these components (not reagents though), and $500 gel box + power supply products are available commercially. But not for the gel imager.

So! Here at genefoo we're working on an affordable & open-source design everything needed to do gel imaging: a transilluminator + geldoc & camera. Here's the plan:

transilluminator

Blue led transilluminator that couples to imaging hood. Big - 9 x 12 in? Possibly with custom manufactured acrylic filters. Built with LED strips (5050SMDs?) One button: on / off. How is the chassis assembled?

hood

Light-tight gel imaging hood that holds camera module, enables z-movement for focus / zoom, perhaps holds filter for camera module, and mates with the top of blue transilluminator. Vacuum cast.

imager + server

Low-cost SoC server + camera - possibly raspberry pi + raspicam module or beaglebone + usb camera, a la openrov.org.

Server (implemented in node?) provides on-demand low-latency low-res ("live") preview of the inside of the geldoc chamber and prompt for capturing a full-resolution image. Image is saved locally and optionally pushed to dropbox (or evernote via email). Images can also be printed with google printer sharing. Supports mobile and desktop browsers.

We're working on a "full-stack* gel imaging system. Along the way I've reviewed some of the available components for existing systems. Here is a list of some common transilluminators.

Transilluminators

UVP Visi-Blue™ Transilluminator - $1360

Life Technologies Safe Imager - $1333

Clare Chemical Dark Reader - $475

Vernier BlueView Transilluminator - $419

Syngene UltraSlim LED - $395

Pearl Biotech Blue Light Transilluminator - $299

IO Rodeo UV Transilluminator - $265

IO Rodeo Blue Light Transilluminator - $115

Gel Documentation Systems

BioRad Gel Doc™ EZ System - $8500

E-Gel® Imager System with Blue Light Base - $6950

UVP DigiDoc-It® Imaging System - $5,459 - $5459

IO Rodeo Smartphone Enclosure - $330

Cartridge gels

Life Technologies E-Gel GO power supply + transilluminator - $613

Life Technologies E-Gel power supply + E-Gel transilluminator - $952

UVP Visi-Blue™ Transilluminator - $1360

price: $1360

area: 21 x 26 cm

dollar / cm2: $2.49

Note: uses UV bulbs and a special filter to downshift the illumination into the blue-light range.

LIFE Technologies Safe Imager - $1333

price: $1333

area: 19 x 19 cm

dollar / cm2: $3.69

Clare Chemical Dark Reader - $475

price: $475

area: 13 x 12 cm

dollar / cm2: $3.04

According to the manufacturer:

The light sources in Dark Reader devices generate maximum light output between 400 and 500 nm - close to where dyes such as SYBR Green, SYPRO Orange, eosin, fluorescein and ethidium bromide are excited.

Sounds like it uses the popular 470 nM LEDs found in some of these other units.

Vernier BlueView Transilluminator - $419

price: $419

area: 11 x 11 cm

dollar / cm2: $3.46

Syngene UltraSlim LED - $395

price: $395

area: 12 x 7 cm

dollar / cm2: $4.70

Two arrays of 54 high intensity 470nm blue LEDs (108 total) illuminate samples from the side. This may provide a better signal to noise ratio than inline illumination.

Pearl Biotech Blue Light Transilluminator - $299

price: $299

area: 10 x 10 cm

dollar / cm2: $2.99

Very simple. Light source may not be uniform.

IO Rodeo UV Transilluminator - $265

price: $265

area: 7.5 x 7.5 cm

dollar / cm2: $4.71

IO Rodeo Blue Light Transilluminator - $115

price: $115

area: 6 x 7 cm

dollar / cm2: $2.74

Small area, may not fit all gel trays. Works w/ IO Rodeo gel box.

Gel Documentation Systems

UVP DigiDoc-It® Imaging System - $5,459

http://www.uvp.com/digidocit.html

E-Gel® Imager System with Blue Light Base - $6950

http://www.lifetechnologies.com/order/catalog/product/4466612

BioRad Gel Doc™ EZ System - $8500

http://www.bio-rad.com/en-us/product/imaging-systems/gel-doc-ez-system

[IO Rodeo Smartphone Enclosure - $330

http://www.iorodeo.com/content/smartphone_enclosure

Sheet-metal case that fits IO Rodeo's UV Transilluminator & Blue Light Transilluminator and secures a smartphone or tablet to the top. $90 filters are an optional accessory to improve the S/N.

Cartridge gels

Life Technologies E-Gel GO power supply + transilluminator - $613

http://www.lifetechnologies.com/order/catalog/product/G4400#

price: $613

area: fits one e-Gel go cassette, 4 lanes

e-Gel cassette price: $6.10

dollar / lane: $$1.53

Life Technologies E-Gel iBase power supply + E-Gel Safe Imager transilluminator - $952

http://www.lifetechnologies.com/order/catalog/product/G6465#

price: $952

area: fits one e-Gel cassette, 12 lanes

e-Gel cassette price: $10.60

dollar / lane: $0.88

E-Gel Safe imager without power supply is $669

Biological protocols are terse, telegraphic recipes that document common wetlab tasks, for instance, sorting DNA molecules by length via gel electrophoresis, growing a liquid culture of E coli, or extracting plasmid DNA from E coli.

Protocols are generally written and shared in plain text with no machine-friendly structure, which is a shame, because all protocols have some elements in common: they specify functionally important inputs, outputs, and a series of biological operations that will transform the inputs into the outputs. Protocols are wetlab functions composed of a series of biological operations with well-defined inputs and outputs.

I've been discussing this in the context of wetlab workflow planning at the diybio google group. Here are some of my thoughts:

A simple way to play with structured protocols would be to establish a small controlled vocabulary for describing the inputs and outputs to various protocols. Meta-protocols could be created by searching for a path from a desired input to a desired output. For example, here is an input and output that would require stitching together several protocols:

Input: 10 ug 1uM plasmid DNA (BBa_I20259) Output: 400 Relative Fluorescence Units, 500nm.

The input / output controlled vocabulary is potentially very complex. I'm not sure how to deal with units.

Looking forward, if the vocabulary of available biological operations (verbs) and inputs/outputs (nouns) was structured in a machine-readable way, it could form the basis of a biological "test-driven development" system. First, desired biological outcomes would be expressed using the vocabulary. Then the biologist would execute an automatically-generated protocol until the tests passed.

Amplicons from three peeps in my fungal barcoding workshop visualized on a gel*. We amplified the 5.8s rRNA subunit using the standard fungal ITS primers.

Patrick used a mildew sample from his shower head in his bathroom, Sara used a Shitake mushroom a local supermarket, and Jesse isn't telling where his fungal sample came from :)

Amplicons are expected to be around 650bp (see this hackpad for more info). Patrick's sample worked best. Patrick & Sara's lanes have smaller bands with a series of smaller amplicons. Perhaps this is because they took surface samples from their mushrooms that were contaminated with other ambient fungi. Perhaps sequencing will be a mess or perhaps it will work... stay tuned.

Startup Weekend: Hadware Edition. Our project: DIY GEL DOC

Magic ingredients for a gel doc: light source, pre-filter, & post-filter. This set works with gelgreen and sybrsafe. Suppliers:

LED matrix: LC2088TB1A from LC-LED.com

pre-filter (blue): Acrylic 2424 Blue from Ridout Plastics