Welcome to my logbook and documentation page ;)

My inspiration comes from Joi Ito, the director of the MIT Media Lab. At the Solid Conference 2015 he gave a talk about the future of technology. His statement is ‘Bio is the New Digital’. His quote on which my project it based is: ‘The design of the future is hardware, it’s software, it’s biology and it’s complex.. But you have to know math.’

My goal of the Biohack Academy is to try and tinker with the future of biotechnology myself.

The design of the future is … hardware

For this part I made a simple but elegant webcam microscope.

The reason is that my focus during the Biohack Academy is on labtime. Nevertheless, the (DIY) hardware is a vital part of the philosophy of both the Waag as well as the quote of Joi Ito. Therefore I decided I wanted to make at least one thing myself.

From all of the gear in the Fablab I especially liked the 3D printer. I used the Prusa printer to make 3 units for the webcam microscope.
This project is done. I even shared the pictures of the magnified cell (with the nucleas and membrames) at a talk.

The design of the future is … biology (part 1)

This was kind of an extra project which I did not expect. But I found a clear instruction video on Youtube how to extract DNA from strawberries.

Because of my fascination with DNA I decided I want to test this protocol for myself. And it worked!

These are the steps:

• But a couple of strawberries in a sealable plastic bag
• Squash the strawberries till it is a mouse
• Pour the strawberries into a cup
• Add one spoon of detergent. The detergent will burst the lipid membrames of the cells and allow the DNA to be extracted
• Mix the strawberries and detergent for at least 30 seconds till little bells arise
• Carefully pour 95% percent alcohol into the glass, near the side of the glass. The alcohol will shed the DNA from the other cellmaterial. The white DNA strengs will float onto the surface.
  This project is done. I even demonstrated this procedure during a talk in which one part was about DNA and genetics.

The design of the future is … biology (part 2)

When I started with Biohack Academy course, I really wanted to do a project with genetic engineering.

My first idea was to work with a CRISPR/cas9 kit, because I am fascinated by the possibilities of gene-editing for agriculture, in healthcare, for industrial use, etc.

Because of European regulations I am not allowed to work with CRISPR/cas9 on living organisms (including myself) at the Waag. A fellow student at the Waag is doing her project about the ins and outs of this regulation, so I am following her insights with a lot of anticipation.

As an alternative I ordered a Biotechnology Explorer kit of Bio-rad, namely the Restriction digestion and analysis of Lambda DNA kit. Because restriction enzymes (RE) are cutting DNA and not cutting and pasting (like CRISPR/cas9), I allowed to work with this experiment at the Waag.

The steps in the proces were 1) restriction digestion, 2) gel electrophoresis and 3) staining DNA. The kit also came with a very insightfull manual. With this manual I got a better understanding of the workings of RE and gel elecrophoresis, but it also gave me another change to practice in the lab. All the steps and details are not that important (and you can look it up on the website of Bio-rad or in Youtube video’s). That is why I want to share all the failures, mistakes and lessons learned :)

Lessons

These are my insights and lessons learned with the RE kit:

• RE were a catalyst for molecular biology. In a way they are the precursors of CRISPR/cas9.
• RE are proteins that cut DNA at specific sites. Hmm, they are not virusses like in CRISPR?
• The DNA and RE need to be kept at minus 20 degrees. Why? Does it otherwise deteriorate?
• The DNA it to cut is of the bacteriophage lambda. It is harmless to mann. Hmm, a bacteriophage is a virus? So virusses can also be cut?
• Gel electrophyses can be compared to a game on the fair, where you put a coin and the top and then it falls down (if you add a couple of coins together the fall down a lot slower).
• Staining DNA pinpoints its location on the gel.
• Doing these kinds of biotechnology also involves a lot of maths.
• In a way it looks like cooking a recipe.

Failures

These are my failures, mistakes and lessons learned with the RE kit:

• Always clear your bench of workplace when you start with an experiment. Yeah, I forget this once.
• Do not be to stingy on the buffer or agarose. Better make too much than not enough.
• Because you work with DNA and RE, you need to work sterile to prevent contamination. I choose to work with gloves (and not add a flame, because the gloves are really flamable)
• Pipeting is also a matter of experience. Remember: soft stop, into the tube, let go, transfer with full press!
• Test if the basket with the tape (in which the agarose becomes solid) is not leaking. Do that in advance!
• Do not pour the gell in the basket when it is still to hot (that is really awfull, because the agarose is really sticky). Wait till you can touch the glass with your hands.
• Do not forget to add dye to the marker DNA, otherwise you do not see a band. Yeah, I did forget that…
  

The design of the future is … software

I want to order a part of a DNA strand at Eurofins Genomics.
First I copied a sample out of Benchling and also tried to work with BLAST and N20. I think it is a great idea to order the lambda DNA like in my RE kit.

Preparation

These were the steps in the preparation:

• I got the sequence here: http://www.bioinformatics.nl/molbi/SCLResources/Sequences.htm#The_sequence_of_lambda_DNA
• I copied the sequence into Benchling.
• You can see the DNA sample here: https://benchling.com/s/seq-UGZfDiXPaX0Tm1LH4Qp5
• First I was not sure how to see the cut sites. This confused me, because the cut sites were there in the RE kit from Bio-rad.
• Roland gave me advice. In the menu you can select Digest (the scissors) and search for the cut sites.
• I wanted to use one or multiple cutsites that were in the kit. These are EcoR1, Pst1 and Hind3.
• A restriction (pun intended) I had was that the synthetic DNA could be only be 500 basepares.
• I searched for a site were the cutsites were close, but I could not find it. Therefore I copied a part (see annotation)
• At Eurofins I copied the sequence, which included a Hind3 cutsite, and added an EcoR1 cutsite at the beginning and an Pst1 cutsite at the end.
• Next I also had to order primers to multiply the DNA in a PCR or thermocycler.
• I had no clue, so I used the PCR Primer Designer of Eurofins https://www.eurofinsgenomics.eu/en/ecom/tools/pcr-primer-design/
• So I copied my DNA sequence into the field. The options I kept at their standard values, like Design Parameters, Primer Criterias, Amplicon Criterias, Reaction Conditions and Output Parameters.
• Next I ordered the synthetic DNA

Execution

These are the next steps:

• Multiply the DNA with the PCR
• Do the RE kit experiments one more time with the Lamda DNA (of the kit) and the synthetic (part of the) Lambda DNA.