1. Synthetic Biology Overview
Seed Magazine –
Synthetic Biology
Download original (large) image

2. Synthetic Biology
Synthetic biology is a new area of biological research that combines science and engineering.
Synthetic biology encompasses a variety of different approaches, methodologies and disciplines, and many different definitions exist.
What they all have in common, however, is that they see synthetic biology as the design and construction of new biological functions and systems not found in nature.

3. Key enabling technologies. DNA sequencing
Synthetic Biology
Key enabling technologies.
DNA sequencing
DNA sequencing is determining the order of the nucleotide bases in a molecule of DNA.
First, large-scale genome sequencing efforts continue to provide a wealth of information on naturally occurring organisms. This information provides a rich substrate from which synthetic biologists can construct parts and devices.
Second, synthetic biologists use sequencing to verify that they fabricated their engineered system as intended. Third, fast, cheap and reliable sequencing can also facilitate rapid detection and identification of synthetic systems and organisms.
Fabrication
A critical limitation in synthetic biology today is the time and effort expended during fabrication of engineered genetic sequences. To speed up the cycle of design, fabrication, testing and redesign, synthetic biology requires more rapid and reliable de novo DNA synthesis and assembly of fragments of DNA, in a process commonly referred to as gene synthesis.

4. History: Synthetic Biology
In 2002 researchers at SUNY Stony Brook succeeded in synthesizing the 7741 base poliovirus genome from its published sequence, producing the first synthetic organism. This took about two years of painstaking work.
In 2003 the 5386 bp genome of the bacteriophage Phi X 174 was assembled in about two weeks.
In 2006, the same team, at the J. Craig Venter Institute, has constructed and patented a synthetic genome of a novel minimal bacterium, Mycoplasma laboratorium and is working on getting it functioning in a living cell.
In 2010, Venter's group announced they had been able to assemble a complete genome of millions of base pairs, insert it into a cell, and cause that cell to start replicating.

5. Synthetic Biology –Craig Ventner 2008
Synthetic Biology –Craig Ventner Complete Chemical Synthesis, Assembly, and Cloning of a Mycoplasma genitalium Genome

6. Complete Chemical Synthesis, Assembly, and Cloning of a Mycoplasma genitalium Genome Daniel G. Gibson, Gwynedd A. Benders, Cynthia Andrews-Pfannkoch, Evgeniya A. Denisova, Holly Baden-Tillson, Jayshree Zaveri, Timothy B. Stockwell, Anushka Brownley, David W. Thomas, Mikkel A. Algire, Chuck Merryman, Lei Young, Vladimir N. Noskov, John I. Glass, J. Craig Venter, Clyde A. Hutchison III and Hamilton O. Smith* Abstract We have synthesized a 582,970–base pair Mycoplasma genitalium genome. This synthetic genome, named M. genitalium JCVI-1.0, contains all the genes of wild-type M. genitalium G37 except MG408, which was disrupted by an antibiotic marker to block pathogenicity and to allow for selection. To identify the genome as synthetic, we inserted “watermarks” at intergenic sites known to tolerate transposon insertions. Overlapping “cassettes” of 5 to 7 kilobases (kb), assembled from chemically synthesized oligonucleotides, were joined by in vitro recombination to produce intermediate assemblies of approximately 24 kb, 72 kb (“1/8 genome”), and 144 kb (“1/4 genome”), which were all cloned as bacterial artificial chromosomes in Escherichia coli..

7. Most of these intermediate clones were sequenced, and clones of all four 1/4 genomes with the correct sequence were identified. The complete synthetic genome was assembled by transformation-associated recombination cloning in the yeast Saccharomyces cerevisiae, then isolated and sequenced. A clone with the correct sequence was identified. The methods described here will be generally useful for constructing large DNA molecules from chemically synthesized pieces and also from combinations of natural and synthetic DNA segments

8. Linear GenomBench (Invitrogen) representation of the circular 582,970-bp M. genitalium JCVI-1.0 genome. Features shown include locations of watermarks and the aminoglycoside resistance marker, viable Tn4001 transposon insertions determined in our 1999 and 2006 studies (3, 4), overlapping synthetic DNA cassettes that comprise the whole genome sequence, 485 M. genitalium protein-coding genes, 43 M. genitalium rRNA, tRNA, and structural RNA genes, and B-series assemblies

9. A plan for the five-stage assembly of the M. genitalium chromosome
A plan for the five-stage assembly of the M. genitalium chromosome. In the first stage of assembly, four cassettes are joined to make an A-series assembly approximately 24 kb in length (assembly contained five cassettes). In the next stage, three A-assemblies are joined together to make a total of eight ∼72-kb B-series assemblies (assembly B62-77 contained four A-series assemblies). The eighth-genome B-assemblies are taken two at a time to make quarter-genome C-series assemblies. These assemblies were all made by in vitro recombination (see Fig. 3) and cloned into E. coli using BAC vectors. Half-genome and whole-genome assemblies were made by in vivo yeast recombination. Assemblies in bold boxes were sequenced to verify their correctness. For the final molecule, the D-series half molecules were not employed. Rather, we assembled the whole molecule from the four C-series quarter molecules.
A plan for the five-stage assembly of the M. genitalium chromosome

10. Assembly of cassettes by in vitro recombination.
Assembly of cassettes by in vitro recombination. (A) Diagram of steps in the in vitro recombination reaction, using the assembly of cassettes 66 to 69 as an example. (B) BAC vector is prepared for the assembly reaction by PCR amplification using primers as illustrated. The linear amplification product, after gel purification, is included in the assembly reaction of (A), such that the desired assembly is circular DNA containing the four cassettes and the BAC DNA as depicted in (C).
Assembly of cassettes by in vitro recombination.

11. Yeast TAR cloning of the complete synthetic genome.

12. Synthetic Biology – Craig Ventner 2010

13. Summary: What is synthetic biology? Synthetic biology refers to both:
the design and fabrication of biological components and systems that do not already exist in the natural world
the re-design and fabrication of existing biological systems.

14. There are two types of synthetic biologists.
Synthetic Biology
There are two types of synthetic biologists.
The first group uses unnatural molecules to mimic natural molecules with the goal of creating artificial life.
The second group uses natural molecules and assembles them into a system that acts unnaturally

15. What is the difference between synthetic biology and systems biology?
Systems biology studies complex biological systems as integrated wholes, using tools of modeling, simulation, and comparison to experiment. The focus tends to be on natural systems, often with some (at least long term) medical significance.
Synthetic biology studies how to build artificial biological systems for engineering applications, using many of the same tools and experimental techniques. But the work is fundamentally an engineering application of biological science, rather than an attempt to do more science.