GeneArt® Algae Engineering Kits contain optimized media for transformation and optimally growing photosynthetic microalgae Chlamydomonas reinhardtii.

GeneArt® Algae Engineering Kits contain optimized media for transformation and optimally growing photosynthetic microalgae Chlamydomonas reinhardtii.

Synthetic Biology lead R&D specialist at Life Technologies in Carlsbad, California, Dr. Farzad Haerizadeh, along with his team, are developing synthetic biology parts, devices and chassis for a variety of industrially relevant organisms including algae, yeast and bacteria.

With a PhD in Genetic Engineering from the University of Tehran, Iran, and another PhD in Molecular Biology from the University of Melbourne, Australia, Dr. Farzad Haerizadeh joined Life Technologies after a stint at Codexis Inc., in Redwood City, CA, where he led projects on metabolic and pathway engineering for production of biofuels and value-added chemicals.

Dr. Haerizadeh has nearly 10 years of experience in industrial microbiology and metabolic engineering, and his expertise on physiology and manipulation of variety of organisms includes algae, plants, yeasts, fungi, and bacteria.

Considering that genetic engineering is one of the most controversial and misunderstood aspects of algal biology, we were interested in getting his take on this rapidly evolving field, what his work is like at Life Technologies, and his observations on the benefits of genetic engineering in the world of algae.

What first got you involved in algal genetic research?

I got involved with algae research when I came to Life Technologies about two years ago. I had been at Codexis Inc. in the Bay Area, where I had worked on metabolic and pathway engineering, mainly in yeast and E. coli, for the production of biofuels and other bio-based compounds with commercial potential.

It’s exciting to be in algae because it’s a young field with enormous, untapped potential. If you think about how long we have been working with other organisms, it’s much longer – plants, for example, have been bred for something like 10,000 years. Then in the 1960s, the introduction of new cropping systems, crop protection and tools brought about the “green revolution,” initiated by Norman Borlaug, which significantly increased the amount of calories produced per acre. The revolution in algae is still ahead of us.

Your team at Life Technology is developing synthetic biology “parts, devices and chassis” for algae and other microorganisms. Please describe that work further.

At Life Technologies, we are developing “tool kits & methods” to make it easier for scientists in both academic and industry labs to work with algae. Right now, there are many different strains being used, so everyone has to invent their methods from scratch. We’d like to make things easier, and to that end, launched our GeneArt® Algae Engineering Kits for Chlamydomonas reinhardtii and Synechococcus elongates. We selected these two organisms to start with because they are frequently used as model systems. Moving forward, we are working to release kits such as ultra transformation reagents for organisms that are currently being used in industrial production such as Chlorella.

The kits contain three items: a frozen strain of algae, growth media that has been optimized for each strain, and a genetic engineering tool kit. Everything is designed to work together and to make molecular biology easy and accessible to microbiologists who may have many years of experience in cultivating algae and other strains but have not worked with genetic techniques before, or scientists that have not worked with algae before, and would like to start or use it in applications such as protein expression. We call our Chlamydomonas “green yeast” and our Synechococcus, “green E. coli,” which means that they can be used as an alternative to yeast and E.coli by people who like to work with CO2 capturing photosynthetic systems.

We also actively develop bioinformatics, including in silico (performed on computer or via computer simulation) models for various algae. These capabilities help our customers to engineer their strains for any end product they want.

Because whatever your application or algae strain you’re working with, you’ll be able to progress much more quickly by genetic engineering of your stock than by altering growth conditions alone. And the techniques are not difficult.

DNA is introduced to algae cells by a number of methods, including biolistic particle delivery system

DNA is introduced to algae cells by a number of methods, including biolistic particle delivery system.

Of your genetic work with algae, what properties have you been working to optimize?

Our work on my group takes place on two fronts. One, we put systems and tools in the hands of customers so that they can start on algae, or optimize strains for themselves. More recently, we have begun to work directly with some industrial customers in a partnership mode, handling the characterization and metabolic genetic engineering of their proprietary algal strains. In the future, we plan to offer this service more broadly on a fee-for-service model.

The properties that our customers seek to optimize are those same traits important to everyone working in the algae field. First, yield – more oil, for instance, for biofuel production, or protein for biotherapeutic production. Algae growers are also highly concerned with contamination, especially in open pool situations. Therefore a good deal of work is going into crop protection. One of our collaborators, Dr. Susan S. Golden, at the University of California, San Diego, recently identified mutations in Synechoccus that confer resistance to certain amoebae. Theoretically, these mutations could be introduced to any strain by genetic engineering techniques to make them resistant.

Can you walk us through the genetic engineering/manipulation workflow? What are the steps and what tools do you provide to facilitate the process?

The workflow is quite streamlined and systematic. The first step is to construct your delivery system. This means that you put the gene of interest, a resistance gene, for example, into what is called a vector, literally a small ring of DNA. Then that vector carrying your gene is introduced into algae cells by a process called transformation – totally similar to E. coli and yeast.

The GeneArt® Algae Engineering Kits contain vectors optimized for Chlamydomonas and Synechococcus, respectively, as well as the tools you need to insert genes into them. You use an enzyme to open the circle, then another to close it back up after insertion. To enable these steps, the kits contain our proprietary TOPO® technology, which has been used by researchers in yeast, bacteria and mammalian cells for many years.

Transforming algae cells is done by mostly electroporation. Transforming algae is still a bit tricky, an issue we are working to tackle right now. Our next versions of GeneArt® kits will contain reagents and a protocol to simplify and boost the efficiency of transformation by hundreds of folds.

How do you compare various algal strains that you have worked with, in terms of their abilities to express the characteristics you are interested in modifying or enhancing?

Something all algae strains have to offer is their rapid growth rate as compared to other organisms used for industrial purposes, especially plants. Some vaccines, for example, are currently produced in plants – a process called “molecular pharming.” But it can take a year to grow and harvest a plant crop, whereas for algae the doubling time is about 8 hours – obviously, you can greatly accelerate timelines and save money by doing things in algae, but the field has a long way to go yet before that is feasible.

Algae colonies successfully transformed with DNA are selected on selective agar plates.

Algae colonies successfully transformed with DNA are selected on selective agar plates.

Where do you see the most potential currently in the research being done on algal biology?

I see the most potential in using genetic tools and engineering techniques to optimize strains. Without that, the field will not progress. Right now, for example, a virus could wipe out thousands of gallons of growing algae. We need to engineer resistant strains, as well as strains with higher yields.

I’m excited about the basic research work we are doing with SD-CAB (San Diego Center for Algae Biotechnology) on industrial strains of algae. We are collaborating on many fronts, such as pathway modeling, tools and technology development. In order to engineer any organism, you need to first characterize it – you need to know how many genes there are, which are functional and how they are connected in pathways. Then we can determine which parts of the pathway we can manipulate – which genes would be beneficial to add, for example.

Tell us about the frozen strains used in the Life Technologies kits

As in being able to freeze and thaw any organism, the problem is in preventing formation of ice crystals. Our method is proprietary, but I can tell you that we put a lot of effort into developing buffers and a complicated freezing protocol that allows the strains in our kits to remain frozen indefinitely.

This offers two key advantages to the end user. First, it gets around the contamination issues that often arise when algal slants are shipped. Second, it eliminates the genetic drift and contamination problem – an immense problem in this field.  I think anyone working with algae knows that the strain you are working with today might not be the same as the one you had six months ago.

Access to frozen stocks means that you can go back to a fresh and original source of cells identical to your starting material. This enables a new level of standardization and uniformity, which we need to move the algae field forward. We are planning to make this protocol available to the community in the near future, meaning they can use our kits to make their own frozen stocks.

If you put your imagination ten years into the future, how would you describe what you think might exist in the world of algal biology?

I hope we will have at least one or two organisms fully characterized and engineer-able for each application, such as biofuels, biotherapeutic production, etc. If we can find a small number of perfect “workhorses,” we will be much better off than we are now with hundreds of strains in use and none of them fully optimized.

If you look at the situation in mammalian cells, there are thousands of cell lines in circulation but only a handful in routine use in bioproduction. That’s because those strains have been determined to be the best, and energy has gone into optimizing both the cells and their growth conditions.

That is where the algae field needs to go if we are to have the same measure of success as other areas of bioproduction and truly realize the potential of these organisms. This is where we are putting our effort.