CT 0729

Genome mutagenesis drives the evolution of organisms. Here, we developed a CRISPR-Cas assisted random mutation (CARM) technique for whole-genome mutagenesis. The method leverages an entirely random gRNA library and SpCas9-NG to randomly damage genomes in a controllable shotgunlike manner that then triggers diverse and abundant mutations via low-fidelity repair. As a proof of principle, CARM was applied to evolve the capacity of Saccharomyces cerevisiae BY4741 to produce β-carotene. After seven rounds of iterative evolution over two months, a β-carotene hyperproducing strain, C7-143, was isolated with a 10.5-fold increase in β-carotene production and 857 diverse genomic mutations that comprised indels, duplications, inversions, and chromosomal rearrangements. Transcriptomic analysis revealed that the expression of 2541 genes of strain C7-143 was significantly altered, suggesting that the metabolic landscape of the strain was deeply reconstructed. In addition, CARM was applied to evolve industrially relevant S. cerevisiae CEN.PK2-1C for S-adenosyl-L-methionine production, which was increased 2.28 times after just one round. Thus, CARM can contribute to increasing genetic diversity to identify new phenotypes that could further be investigated by reverse engineering.

For people with disabilities, chances to find or keep work are negatively affected by multiple problems like lower education, poverty and poor health. Furthermore, although active labour market policies proved to be effective for unemployed in general, success rates are poor for persons who are unemployed due to multiple problems. The present study aims to describe the development of a method as well as professional training to teach its application, and to assess the feasibility of method and training. The Strength-based method (CARm) aims to promote employment of work-disability benefit recipients with multiple problems.

The main principles of the Strength model were redesigned for better applicability in a population of work-disability beneficiaries, resulting in the CARm method. As part of the CARm method, a training module for Labour Experts (LEs) was developed. To assess the new designed method and training, a one-group, pre-post design was used. Data were collected from eight participating LEs, five female and 3 male, aged between 41and 55 years and having 2–17 years working experience. We used self-report questionnaires and a semi-structured discussion meeting after the training sessions with the LEs.

Eight labour experts (LEs) from the Dutch Social Security Institute participated in the study. Most LEs felt an improvement in their ability to ascertain developmental needs, opportunities and threats in the client’s situation. Three months after the training, LEs almost unanimously agreed on the statements ‘I expect to use the CARm method more frequently in the future’ and ‘I use the CARm method in daily practice whenever possible’. The overall rating for the training on a scale from 1 to 10 was 7.6 (range 7–9). The overall satisfaction with the trainers was good.

The CARm method and training was found to be a feasible approach to facilitate LEs working at the UWV reintegration service to support clients with multiple problems. Sufficient managerial support for participating LEs is a key factor for successful implementation of CARm. Results show that CARm is worth testing for efficacy in a future trial.