Papers of the Week 12th February 2018

Welcome to our pick of papers and articles from last week. These are a collection of the synthetic biology papers that have captured our attention and that we think you should know about. Maybe you’ve seen some others you’d like to tell us about. Send us a comment or tweet us at @synbiobydesign. See our previous recommended papers here.

CRISPR-Cas – the new Sherlock

A new and improved diagnostic tool, aptly named SHERLOCK, allows tumour cell detection and pathogen detection via RNA and DNA signals. This CRISPR-Cas13-based system gives simple colorimetric readouts via a paper test. Once the molecule in question is detected, the enzyme cuts additional RNA, releasing a fluorescent dye. SHERLOCK has been developed so that it can perform multiple tests simultaneously, such as distinguishing between Zika and Dengue virus infections in a sample.

Detection of viral ssRNA, similar to a pregnancy test, via the new SHERLOCK system. Image from the MIT news article

CRISPR-Cas9 as an event recorder

Another addition to the myriad of CRISPR-cas applications is “cellular recorder”. By harnessing the hugely sensitive CRISPR device dubbed “CAMERA1”, researchers recorded the amount and duration of E. coli exposure to antibiotic. A different recorder, CAMERA2, uses a modified Cas9 enzyme that edits single nucleotides rather cutting DNA. This was used to sequentially record environmental cues such as virus infection and light exposure, and was also implemented in human cells.

Orthogonal ribosomes to improve metabolic flux

Synthetic circuits fail to express proteins to desired levels for a number of different reasons, including competition for the host’s ribosome pool. One way around this is the expression of orthogonal ribosomes with a synthetic 16S rRNA that only translate circuit-specific mRNAs. Circuit genes that were previously coupled due to sharing of the original ribosome pool can then be independently translated, reducing the disconnect between transcription and translation rates. Additionally, Darlington et al.  modelled and built in E. coli a negative feedback controller to direct resources to synthetic circuit gene expression according to demand.

Dynamic allocation of ribosomes. When there is no circuit present (left), most ribosomes are original host ribosomes, and the few orthogonal ribosomes translate the controller (yellow) which then acts to repress the synthesis of the orthogonal 16S rRNA. The introduction of a low demand circuit (centre) results in the orthogonal ribosomes synthesising circuit genes rather than the controller. This increases the pool of orthogonal ribosomes. The introduction of a high demand circuit (right) further decreases the repression of orthogonal rRNA, increasing the amount of orthogonal ribosomes allocated from the host pool. This reduces coupling of the circuit GFP and RFP genes. Image from Darlington et al., Nature Communications 2018.

Gene synthesis in droplets: Dropsynth

Dropsynth is a low cost method for gene synthesis in a water-in-oil emulsion. The genes are built up from a large pool of oligomers, where only the relevant oligos get pulled down into a microbead using barcodes. While still relatively inefficient, Dropsynth is nevertheless an effective tool for library building (at less than $2 per gene), allowing the researchers to build a library of synthetic genes and test their ability to complement E. coli knockouts.




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