Genome editing in the mushroom-forming basidiomycete Coprinopsis cinerea, optimized by a high-throughput transformation system

Abstract The precision and ease of use of CRISPR nucleases, such as Cas9 and Cpf1, for plant genome editing has the potential to accelerate a wide range of applications for crop improvement. For upstream research on gene discovery and validation, rapid gene knock-outs can enable testing of single genes and multi-gene families for functional effects. Large chromosomal deletions can test the function of tandem gene arrays and assist with positional cloning of QTLs by helping to narrow down the target region. Nuclease-deactivated Cas9 fusion proteins with transcriptional activators and repressors can be used to up and down-regulate gene expression. Even more promising, gene insertions and allele replacements can provide the opportunity to rapidly test the effects of different alleles at key loci in the same genetic background, providing a more precise alternative to marker-assisted backcrossing. Recently, Texas A&M AgriLife Research has supported the development of a Crop Genome Editing Lab at Texas A&M working towards optimizing a high-throughput gene editing pipeline and providing an efficient and cost-effective gene editing service for research and breeding groups. The lab is using rice as a model to test and optimize new approaches aimed towards overcoming current bottlenecks. For example, a wealth of genomics data from the rice community enables the development of novel approaches to predict which genes and target modifications may be most beneficial for crop improvement, taking advantage of known major genes, high-resolution GWAS data, multiple high-quality reference genomes, transcriptomics data, and resequencing data from the 3,000 Rice Genomes Project. Current projects have now expanded to work across multiple crops to provide breeding and research groups with a rapid gene editing pipeline to test candidate genes in their programs, with the ultimate goal of developing nutritious, high-yielding, stress-tolerant crops for the future.

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Shaping the nebulous enhancer in the era of high-throughput assays and genome editing

Briefings in Bioinformatics ◽

10.1093/bib/bbz030 ◽

2019 ◽

Vol 21 (3) ◽

pp. 836-850

Author(s):

Edwin Yu-Kiu Ho ◽

Qin Cao ◽

Mengting Gu ◽

Ricky Wai-Lun Chan ◽

Qiong Wu ◽

...

Keyword(s):

Genome Editing ◽

High Throughput ◽

Wide Spectrum ◽

Data Interpretation ◽

Future Studies ◽

Transcriptional Enhancers ◽

The Past ◽

Genome Wide ◽

Gray Areas ◽

Practical Implications

Abstract Since the 1st discovery of transcriptional enhancers in 1981, their textbook definition has remained largely unchanged in the past 37 years. With the emergence of high-throughput assays and genome editing, which are switching the paradigm from bottom-up discovery and testing of individual enhancers to top-down profiling of enhancer activities genome-wide, it has become increasingly evidenced that this classical definition has left substantial gray areas in different aspects. Here we survey a representative set of recent research articles and report the definitions of enhancers they have adopted. The results reveal that a wide spectrum of definitions is used usually without the definition stated explicitly, which could lead to difficulties in data interpretation and downstream analyses. Based on these findings, we discuss the practical implications and suggestions for future studies.

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High-throughput Genome Editing Tools for Lactic Acid Bacteria: Opportunities for Food, Feed, Pharma and Biotech

10.20944/preprints201809.0354.v1 ◽

2018 ◽

Cited By ~ 1

Author(s):

Rosa Aragão Börner ◽

Vijayalakshmi Kandasamy ◽

Amalie Melton Axelsen ◽

Alex Toftgaard Nielsen ◽

Elleke F Bosma

Keyword(s):

Lactic Acid ◽

Lactic Acid Bacteria ◽

Genome Editing ◽

High Throughput ◽

Recombinant Dna ◽

Cell Factory ◽

Rapid Construction ◽

Design Build ◽

Recombinant Dna Techniques ◽

New Applications

This mini-review provides an overview of traditional, emerging, and future applications of lactic acid bacteria (LAB) and discusses how genome editing tools can be used to overcome current challenges in all these applications. It also describes currently available tools and how these can be further developed, and takes current legislation into account. Genome editing tools are necessary for the construction of strains for new applications and products, but can also play a crucial role in traditional ones, such as food and probiotics, as a research tool for understanding mechanistic insights and discovering new properties. Traditionally, recombinant DNA techniques for LAB have strongly focused on being food-grade, but they lack throughput and the number of genetically tractable strains is still rather limited. Further tool development in this direction will enable rapid construction of multiple mutants or mutant libraries on a genomic level in a wide variety of LAB strains. We also propose an iterative Design-Build-Test-Learn workflow cycle for LAB cell factory development based on systems biology, with “cell factory” expanding beyond its traditional meaning of production strains and making use of high-throughput genome editing tools to advance LAB understanding, applications and strain development.

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