How to start learning about DIY BIO

I am completely self taught in Genetic Engineering/Synthetic Biology. I became fascinated with Synthetic Biology after the initial success of the crowdfunded project Glowing Plant on Kickstarter. Glowing Plant was ultimately a failure, because the founder of the project was not familiar enough with the industry and, I believe due to that, misplaced much of the money he raised (over $750,000!).

For beginners the best place to start is online:

(1) EDX.org

By far the best online resource is EDX.org

The biology courses on edx.org are extensive and include basic biochemistry, to genetics, industrial biotechnology and even a course on synthetic biology. Personally, I would start with MITx’s course Quantitative Biology workshop and the introductory courses on Biochemistry from RICEx as well as the BioChemistry course from Harvardx. So far, the best intro class to genetic engineering I have found is “Making Biologic Medicines for Patients: The Principles of Biopharmaceutical Manufacturing” by MITx on edx. Good luck and as the website changes I may need to update the aforementioned list.  EDX is amazing, I have taken courses ranging from Drug comercialization, to Biochemistry, to Genetics.

(2) Coursera.org

Coursera is another excellent resource, although they are not as extensive as EDX. They had some very good courses on Virology on their website as well as some information on genetics, be sure to go to their Life Sciences catalog.

(3) Futurelearn.com

Futurelearn is another one, I have taken a course or two there, definately one to check occasionally for new content.

All the courses I mentioned are free, now initially, I had it in my head that I would pay for the courses to get certified, but I have found that no one really cares about online courses. That is why it is important to document your work online in social media, like on blogs such as this, Youtube, Instagram and so on.

(4) Books

• Recombinant DNA: Genes and Genomes – A Short Course
• Molecular Genetics of Bacteria

Excellent book that I have learned much from, after building a foundation in some online courses that I mentioned above.

 

 

The Holy Grail: AgroBacteria License APHIS and BRS permit

The Holy Grail of plant, genetic Engineering: De-weaponized Agrobacteria, that is with its oncogenes removed, is considered a genetically modified organism and is regulated by the USDA, when it crosses state lines.

Agrobacteria is a rather commonly used vector for genetic engineering. This whole process has been rather difficult, so I decided to add this blog to help others who may be thinking of getting into genetic engineering of plants.

In order to get Agrobacteria you have to go through a relatively lengthy process first by getting an  APHIS level 2 account, which means you have to personally show up at a registered USDA office and show them your identification, so they know who you are.

After several emails I recieved this via the Biotechnology Regulatory Services (BRS):

“Thank you for contacting APHIS’ Biotechnology Regulatory Services (BRS).  BRS regulates genetically engineered organisms (GE) that may pose a pest risk to plants. If the Agro is genetically engineered, you will need a BRS permit.  You can learn more and apply online at:  https://www.aphis.usda.gov/aphis/ourfocus/biotechnology/permits-notifications-petitions. For additional information, please see the Permit User’s Guide at: https://www.aphis.usda.gov/biotechnology/downloads/permit_guidance.pdf.”

Now the Permit User’s Guide is helpful and has many examples in the appendix.

I was confused as to after I had applied for a BRS permit, if I needed them to inspect my organization (Dallas Maker Space) or not, their answer:

“You will need just the BRS permit. Once you submit the permit application with your data, our biotechnologists will determine whether you need an inspection and provide that information to you.”

Before you start filling out the BRS form in epermit website, you need

(1) to have an organization, its address and person of responsibility who is going to send you the bacteria;

(2) you will also need to be able to define the genes and vectors you intend on using.

Once you have made the necessary contacts, go to here to start your account.

Once you log in click Create\Renew\Amend

a 

If you don’t finish it the first time (you won’t) you can continue by clicking “My Application”

Click this link to get a document I created to help you get through the APHIS BRS permit appliation, it is a step by step screen shot of the process.

Thanks to Cory Tobin, Colleen Wood of BRS and the DIYBIO community for their help.

Teaching My First Class on Genetic Engineering/Synthetic Biology

classWe had a large group of people come for our first Genetic Engineering/Synthetic Biology at Dallas, it was held at Dallas Maker Space, we started by preparing for the class several days in advance by prepping the MS medium for the Tissue Culture Lab.

The Tissue culture was Multi solution, from Microclone.

22 in attendance to yesterday’s first SynBio class.
5 first time guests
2 new DMS members Great job with the DMS tour.

Now there was also a lecture before the lab, on the basics of Genetic Engineering

Lecture Slides

https://drive.google.com/file/d/0BxnJkKTcTEksSGJKUkY1Qk9zMUk/view?usp=sharing

Plants From Test Tubes An Introduction to Micro-Propogation 4th Edition book review

I bought this book on the recommendation of several people, and basically this is by far the most well known, modern book on the topic. It covers the history, business, laboratory setup and design, media selection, sterile technique and even a short section on Genetic Engineering with Gene Gun and/or Agrobacteria.

 

Frankly, all of the above information can be had rather easily online and it isn’t in depth enough to be helpful if you are really looking to pursue this as a business, but the real value of the book comes in its kind of encyclopedic listing of media choices for plant species, this is very helpful and can be a guide to the budding Tissue Culture-ist as they venture out. I recommend this book for people, like me, who are starting out in Tissue Culture and who need a guide or Bible to help them illuminate some of the uncertainty as they get started. Well worth it.

AgroBacteria, the BigFoot of DIY BIO

As someone starting out in DIY BIO I have found trying to procure AgroBacteria, to be a living hell. I have been rejected by several instate universities, with a terse warning, the people who are willing to give me the bacteria are out of state. Now according to the USDA since AgroBacteria is de-weaponized, which means the virulent parts have been removed, thus the organism is genetically modified and cannot be imported or sent across state lines without a special form. First an APHIS level 2 account is required, which means you have to go the the nearest office and show them your ID so they can put a responsible party with the account, second you have to fill out a PPQ 526 form, but the way to fill out the form online doesn’t make sense to me, which has me waiting for a convenient time to call. Unfortunately, I work night shift at the moment and have trouble going on for a regular 9-5 schedule. I will update this post as I figure out what to do.

Plant Tissue Culture Microclone Kit Issues/Review

I found that some of the longer containers that are included in the MicroClone kit, the plastic is too brittle and when you push it into the container, they crack and break, nearly all of the containers were cracked, and contaminated with fungis and had to be thrown away.

I still think that the kit, is a good starter kit, it helped me overcome the fear and uncertainty around Tissue Culture, I now know the basics and can move on to more advanced techniques if I so choose.

My Experience at GenSpace Biohacker Bootcamp

I was very determined to go to GenSpace Biohacker Bootcamp in Brooklyn, NY; I live in Dallas, TX and originally was going to go in January, but there was a massive snow storm at that time, that forced me to go in the summer. I made a trip of it, as it was a week long course I stayed at an AirBnB in Brooklyn and explored the city during the day, and learnt about some of the lab techniques at night. It was very enjoyable, I took several guided tours of the city, went to many famous landmarks, worked out at a gym and still had time for the course, each evening.

Dr. Mike Flanagan, was our class instructor, there was an interesting group of people, one was a highschool student interested in creating leather from Synthetic Biology, another was an entrepreneur curious about a budding field of Technology, programmers and PhD students.

I found the class to be well paced for people unfamiliar with Biology or Genetic Engineering. I have been studying Genetics, Synthetic Biology, BioChemistry online for some time, as I have no formal training, I have found EDX.org and Coursera.org to be extremely valuable in this regard. Since I have little lab experience I was very keen on coming and learning as much as I could, and I wasn’t disappointed.

We started by extracting our own DNA with a q-tip and sending it in for analysis, learning the procedure for PCR and Gel Electrophoresis.  We used PCR to amplify a segment of our own DNA which was also used in the Gel Eletrophoresis experiment, the assay ran with a DNA standard as well as 5 samples, since there were 5 students in the class.

 

On the following day, we used restriction enzymes to splice a bacterial plasmid with a antibiotic selection marker and a fluorescent protein, the plasmid was then added to a competent bacteria and set over night in an incubator. The next day we looked at our control and experimental samples to see if we were successful, we were.

We also discussed various Synthetic Biology techniques, opportunities, IGEM, and the future of the field.

On the final day, Dr. Flanagan also discussed our ancestry, on the first day when we had extracted our DNA, some of it was also sent out to a local company for analysis. We discussed the implications as well as possibilities for DIY biohacking.

I recommend the course to anyone who is new to Synthetic Biology and found the people and environment to be very rewarding. I am glad I went.

 

First attempt at Plant Tissue Culture

 

At Dallas Maker Space, we worked to try to use Plant Tissue Culture, we used the MicroClone Starter Kit. The kit is probably a good introduction to plant tissue culture, but because we were new to this, we didn’t know that different types of plants require different media, Orchid is particularly picky and actually can poison itself with its own blood.

The MicroClone kit comes with three types of agar starters, plants themselves have hormones that tell them when to root, or branch, it was found that these hormones can be used by us to have plants root or branch on command -a very useful tool, when cloning plants. The kit comes with a BA hormone for multiplication/branching and also the NAA hormone for rooting/growth. We started with the Multiplication branching media. Following the instructions, we microwaved some Distilled water, poured the agar and Hormone into it as well as added sugar, a necessary ingredient. Then we used our autoclave to sterilize all of the containers, with the previously poured agar in them, I carefully closed the lids, and yet kept them slightly open so that they will not pop under the pressure, I also put my tweezers and blade into the autoclave as well.

Orchid

After 3 weeks of waiting, we found that the Orchid gives a black color in the multi medium, after asking some more experiences BioHackers about this they told us this is the Orchid blood and it will poison them eventually, Orchid media typically contains activated charcoal as well as banana, – who knew.

Fungis

The above picture is of a rose stem, now the stem is still green and alive, but the fungis is over taking the area around it, perhaps it was not perfectly sterilized before being put into the container.

The above picture is of two Rose stems, they are not contanminated with fungis, and will wait for a detailed analysis later.

Lessons Learned:

(1) We washed all of the various plants Orchid and Rose together, this was a mistake and could have led to cross contamination

(2) We needed to Autoclave the cutting board, which we did not do and was another source of contamination

(3) Need special Media for Orchids and need to research proper media for each plant type before moving forward.