Biofoundry-assisted expression and characterization of plant proteins.

Many goals in synthetic biology, including the elucidation and refactoring of biosynthetic pathways and the engineering of regulatory circuits and networks, require knowledge of protein function. In plants, the prevalence of large gene families means it can be particularly challenging to link specific functions to individual proteins. However, protein characterization has remained a technical bottleneck, often requiring significant effort to optimize expression and purification protocols.

Measurement of Transgene Copy Number in Plants Using Droplet Digital PCR.

Transgenic plants are produced both to investigate gene function and to confer desirable traits into crops. Transgene copy number is known to influence expression levels, and consequently, phenotypes. Similarly, knowledge of transgene zygosity is desirable for making quantitative assessments of phenotype and tracking the inheritance of transgenes in progeny generations.

Rational design of minimal synthetic promoters for plants.

Promoters serve a critical role in establishing baseline transcriptional capacity through the recruitment of proteins, including transcription factors. Previously, a paucity of data for cis-regulatory elements in plants meant that it was challenging to determine which sequence elements in plant promoter sequences contributed to transcriptional function. In this study, we have identified functional elements in the promoters of plant genes and plant pathogens that utilize plant transcriptional machinery for gene expression.

Phytobricks: Manual and Automated Assembly of Constructs for Engineering Plants.

Phytobricks are standardized DNA parts for plants that can be assembled hierarchically into transcriptional units and, subsequently, into multigene constructs. Phytobricks each contain the sequences of one or more functional elements that comprise eukaryotic transcription units, with sequence features that enable them to be used interchangeably in one-step cloning reactions to facilitate combinatorial assembly. The simplicity and efficiency of this one-step reaction has enabled Phytobrick assembly to be miniaturized and automated on liquid handing platforms.

Two proteases with caspase-3-like activity, cathepsin B and proteasome, antagonistically control ER-stress-induced programmed cell death in Arabidopsis.

Programmed cell death (PCD) induced by endoplasmic reticulum (ER) stress is implicated in various plant physiological processes, yet its mechanism is still elusive. An activation of caspase-3-like enzymatic activity was clearly demonstrated but the role of the two known plant proteases with caspase-3-like activity, cathepsin B and proteasome subunit PBA1, remains to be established. Both genetic downregulation and chemical inhibition were used to investigate the function of cathepsin B and PBA1 in ER-stress-induced PCD (ERSID).

Endoplasmic reticulum stress-induced PCD and caspase-like activities involved.

Plant cells, like cells from other kingdoms, have the ability to self-destruct in a genetically controlled manner. This process is defined as Programmed cell death (PCD). PCD can be triggered by various stimuli in plants including by endoplasmic reticulum (ER) stress.

The Arabidopsis peptide kiss of death is an inducer of programmed cell death.

Programmed cell death (PCD) has a key role in defence and development of all multicellular organisms. In plants, there is a large gap in our knowledge of the molecular machinery involved at the various stages of PCD, especially the early steps. Here, we identify kiss of death (KOD) encoding a 25-amino-acid peptide that activates a PCD pathway in Arabidopsis thaliana.