Engineered Microbes Clean Up Copper & DNA Cloning Goes Full-Auto (#17)

Plus: CRISPR enables super accurate insertion of large DNA chunks into genomes.

☀️ Good morning.

This Thanksgiving, I’m grateful to you, my readers. In the convoluted chaos of the internet, I know that you could be reading many things. Thank you for reading this.

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DNA Cloning Goes Full-Auto

It’s midnight in the lab. All your co-workers have gone home. You’re alone at the bench, fearful that a security guard will arrive at any moment and kick you out. If you, like me, have found yourself in this situation, it may be because you have failed—repeatedly—to clone a particularly tricky DNA sequence.

But that could happen a lot less in the near future. Robots are coming online, promising to automate entire experiments. So, too, could robots be used to create and clone custom DNA sequences. Unfortunately, a specific step in the cloning process—blasting cells with electricity to coax them into taking up DNA—has proven troublesome for machines.

A new study, published Nov. 24 in ACS Synthetic Biology, offers a fully automated protocol for DNA cloning that relies on “natural transformation”, rather than electroporation. Sang Yup Lee’s lab, at the Korea Advanced Institute of Science and Technology, demonstrated that Acinetobacter baylyi, a type of bacteria, efficiently take up DNA, without the need for specialized equipment. Researchers already knew that A. baylyi were “naturally competent”, but I think this is the first time that these bacteria have been incorporated into a fully automatic, DNA cloning pipeline.

The team simply dropped DNA sequences into liquid with A. baylyi, and the cells gobbled them up and began to reproduce, making tens, then hundreds, then thousands of copies of the DNA.

“No DNA purification, competence induction, or special equipment is required,” the authors write. “Up to 10,000 colonies were obtained per microgram of DNA, while the number of false positive colonies was low.” The team used the new protocol to clone 21 biosynthetic gene clusters, with lengths ranging from 1.5 to 19 kb, and showed that the protocol was relatively consistent when performed by an Opentrons robot. Check out the paper! Link

Engineered Bacteria Clean Up Copper

A team of researchers at the University of York and Umeå University, in Sweden, have engineered E. coli bacteria to accumulate heaps of copper.

To do that, the team fused seven different snippets of proteins—each thought to bind copper ions—to a protein called Maltose Binding Protein, or MBP. These protein chimeras, when expressed inside of the bacterial cells, “conferred tolerance to high concentrations of copper sulphate,” write the authors. The tolerance to copper was so high, in fact, that some bacteria could withstand concentrations “160-fold higher than the recognised EC50 toxic levels of copper in soils.”

The researchers also crunched some data, on computers, and found that these copper-binding proteins might be able to bind other types of metals. The authors suggest that these bacteria could “be adapted for the removal of other hazardous heavy metals or the bio-mining of rare metals.” Let’s mine some asteroids with juiced up space bacteria!

This work was published in Scientific Reports, and is open access. Link

Knocked Down Genes Reveal How Metabolism Heals

Drop a bacterium into a new environment—one with more sugar, or different competitors—and its metabolism will quickly adapt. In fact, the metabolism of a bacterium can change so much, when placed in a new environment, that the total mass of its various enzymes can double, according to a 2016 study by Schmidt et al. in Nature Biotechnology.

Okay, so lots of protein levels change when a cell feels stressed out. But what happens when just one gene, encoding one enzyme, is repressed, or knocked down? How does the rest of the cell’s intricate metabolism shift in response to account for that single, “deficient” gene?

It turns out that cells shift their cellular resources in intriguing ways to “heal” a broken link in the metabolism chain. A new study, published Nov. 24 in Cell Systems, used CRISPRi (CRISPR interference) to repress one gene at a time in E. coli cells. Researchers used an inducible version of CRISPRi that can be turned ‘on’ at will "to investigate how E. coli metabolism responds to decreases of enzyme levels.”

For the study, the team repressed 1,515 different genes involved in metabolism, creating 4-6 sgRNAs for each gene; that resulted in a total of 7,177 unique strains of E. coli. They then “induced” the CRISPRi system, and measured “the time delay between inducer addition and appearance of fitness defects.”

30 of the strains were studied in greater detail, unveiling some interesting examples of metabolisms “adjusting” to account for a deficient enzyme in a pathway. “Overall, our results highlight the central role of regulatory metabolites in maintaining robustness against ever-changing concentrations of enzymes in a cell,” the authors wrote.

To learn a bit more about this study (in general language), check out the press release. This study is open access. Link

Bacteria Evolved in Lab to Study Antibacterial Resistance

Drug-resistant bacteria are a growing issue, and researchers are racing to develop new antibiotics to fight back. To make better antibacterial compounds, scientists must first understand how antibacterial resistance emerges.

Published Nov. 24 in Nature Communications, a team—from RIKEN and the University of Tokyo—used an automated, robotic system to repeatedly passage bacteria over more than 250 generations, while grown in the presence of 95 different antibacterial chemicals. In other words, they put cells into evolutionary overdrive.

The researchers analyzed the evolved cells, studying how gene expression profiles changed for each strain, and then fed their massive dataset into a machine learning algorithm. They identified several gene expression “signatures” that were associated with drug resistance in the microbes. Read the press release on this open access study. Link

Super Accurate Insertion of DNA Chunks with CRISPRi

A special type of CRISPR-Cas system, harnessed from Vibrio cholerae bacteria, enabled researchers to integrate large chunks of DNA, up to 10,000 bases in length, at specific sites in the genome of bacteria. The method is supposedly 100 percent efficient.

The team, from Columbia University, call their method INTEGRATE (insertion of transposable elements by guide RNA–assisted targeting). After demonstrating their method by inserting DNA at one location in the genome, they scaled up their work in a massive way: By expressing multiple guide RNAs in the cells, the team was able to insert DNA chunks at three places in the genome at once. This work was published in Nature Biotechnology. Link

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🧫 Rapid-Fire Highlights

More research & reviews worth your time

  • Did you eat sweet potatoes on Thanksgiving? Maybe next year, they’ll be genetically-engineered. A new study reports a CRISPR-Cas9 based method to edit yam genomes from the Dioscorea family. Plant Biotechnology Journal. Link

  • Luciferase—a group of enzymes that produce a firefly’s glow—has a rich, 160 million year history. A new study, that analyzed Mycena mushrooms, unravels the evolutionary lineage of this “bright” family of genes. PNAS (Open Access). Link

  • Researchers at the University of York identified a gene, in wheat, linked to plant growth. By ‘boosting’ that gene, a modified wheat variety can produce about 12% more food compared to conventional wheat. New Phytologist (Open Access). Link

  • A 3D-printed, open source microscope, costing a few hundred dollars, can produce images that allegedly rival “commercial microscopes that cost up to a thousand times more.” That’s according to a press release from the Leibniz-Institute of Photonic Technology for this new, intriguing study. Nature Communications (Open Access). Link

  • Ribosomes are not the only cellular machines that can make proteins. A new study reports nonribosomal peptide synthesis using DNA as a template. That’s a major achievement; check this study out. Cell Chemical Biology. Link

  • To engineer plant genomes, many researchers use Agrobacterium tumefaciens mediated gene transfer, which leaves behind “foreign DNA” that could lower the efficiency of gene delivery. A new study analyzes two alternative strategies to reduce the amount of foreign DNA left behind. Scientific Reports (Open Access). Link

  • ‘Designer proteins’ are a hot topic right now. Engineered versions of enzymes can withstand higher temperatures, or catalyze reactions at a faster rate. Intriguingly, a new study shows that, even if you modify an enzyme to pack loads of valine amino acids into its interior, the “designer” protein still has “high stability (Tm = 106 °C) even with its reduced and loosened core packing.” PNAS (Open Access). Link

  • Sci-Fi horror flicks often show an engineered microbe escaping the laboratory and wreaking havoc on global populations. That scenario is a bit far-fetched, but it doesn’t mean scientists aren’t concerned about biocontainment, or keeping genetically-engineered bugs in the lab. A new study shows that microbes can be engineered to only grow on solid media by using some fancy chemistry and unnatural amino acids. ACS Synthetic Biology (Open Access). Link

  • A massive new study used CRISPRi to knock down 88 genes in E. coli (using 5,927 different guide RNAs), demonstrating that, if you slowly add mutations to the guide RNAs to change how efficiently they can recognize DNA, you can effectively tune the growth rate of bacterial cells. Nucleic Acids Research (Open Access). Link

  • A new review takes a look at the clinical therapies emanating from synthetic biology research. Synthetic Biology (Open Access). Link

  • By embedding photosynthetic cells in a lipid capsule, researchers created “multicellular spheroids capable of both aerobic (oxygen producing) and hypoxic (hydrogen producing) photosynthesis in daylight under air.” Basically, miniature algal bioreactors. Nature Communications (Open Access). Link

  • By encapsulating various enzymes inside of a viral particle, researchers created nanoreactors that could catalyze the entire glutathione synthesis pathway. ACS Synthetic Biology. Link

  • The famous repressilator genetic circuit—first created by Elowitz and Leibler in 2000—has been recreated, albeit with a programmable, dCas9 protein. ACS Synthetic Biology. Link

  • Engineered E. coli bacteria were embedded into cellulose biomaterials. They may be useful, one day, for real-world biosensing. bioRxiv (Open Access). Link

  • DNA ‘sponges’—which are basically decoy DNA binding sites that compete for a protein’s attention—can be used to tune gene expression in bacteria. Nature Communications (Open Access). Link

  • For a new study, researchers randomly inserted three genes, involved in sucrose utilization, into the E. coli genome. That random insertion produced many different, engineered bacteria that could each grow on sucrose at various rates. ACS Synthetic Biology. Link

  • Metabolisms are messy and complicated. Machine learning can help rewire them in a more predictable way. That’s the focus of this new review. Metabolic Engineering. Link

  • Using a deactivated Cas12a protein, researchers developed a CRISPR system that can both activate, and repress, multiple genes at once in various bacterial species. ACS Synthetic Biology. Link

  • Are you “rewiring” yeast metabolisms to produce, say, beer with a citrus-y flavor, or bread that rises faster? Now, you can engineer your yeast with light! A new study reports a light-controlled method to control gene expression in S. cerevisiae yeast. The authors say that this method can activate “gene expression in only 0.6 h after switching cells from light to darkness…” ACS Synthetic Biology. Link

  • Yeast cells are chock full of proteins and macromolecules. But they are surrounded by a thick cell wall. Researchers placed yeast in water and blasted them with electricity, forcing the cells to release about 90% of free amino acids after 2 hours. Frontiers in Bioengineering and Biotechnology. Link

  • The fusion of synthetic biology and materials science has enabled functional, living materials—think bacterial cells embedded in cellulose, or the “lab coat of the future” from MIT that could detect and analyze would-be chemical hazards. A new review takes a closer look. Trends in Biotechnology. Link


📰 #SynBio in the News

  • Ginkgo Bioworks received a $1.1 billion loan from the U.S. International Development Finance Corp to scale up their efforts in COVID-19 vaccine manufacturing. Reuters. Link

  • Researchers are editing the CCR5 gene in monkeys to potentially treat HIV. The monkeys haven’t yet been exposed to the virus. CCR5 is the same gene edited by He Jiankui during the CRISPR baby scandal. Future Human. Link

  • Biomanufacturing company, Genomatica, has inked a deal with Aquafil, a nylon manufacturing company, to create “a new demonstration scale facility.” Are pantyhose made from microbes on the horizon? Tech Crunch. Link

  • The new Oxford/AstraZeneca vaccine is up to 90% effective, according to interim data. But a dosing mix-up has allegedly sewn confusion amongst scientists and the public. MIT Technology Review (Link) & Science (Link)

  • The Institute for Protein Design has demonstrated that “designer proteins” can bind to, and disable, a part of SARS-CoV-2, the virus that causes COVID-19. Katherine J. Wu reported a nice feature on their work during the pandemic. The New York Times. Link

  • Spiber, a Japanese biomaterials company, and Goldwin, a sportswear brand, are launching a sweater made from synthetic, protein-based materials. Forbes. Link

  • Many well-deserving synthetic biologists were named as 2020 AAAS Fellows. Check out the full list! Science. Link

  • (Not SynBio) Over 500 years, Leonardo da Vinci’s sketches have accumulated their own, unique collection of microbes. Wired. Link

  • (Not SynBio) It’s great if you like eating Impossible burgers, but please protect your bones! Study suggests that “meat-free diets linked with greater risk of breaking bones.” New Scientist. Link


🐦 Tweets of the Week

Cell Reports released a special issue of papers, featuring research at the intersection of space flight and biology. Check it out! 👇

One of the papers in the special issue specifically caught my eye: a longitudinal study on aging, based on 520 days of simulated space travel.


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