Modern Biotechnologies: Innovative and Sustainable Approaches for the Improvement of Sugarcane Tolerance to Environmental Stresses

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Abstract

Sugarcane (Saccharum spp.) is one of the most important industrial cash crops, contributing to the world sugar industry and biofuel production. It has been cultivated and improved from prehistoric times through natural selection and conventional breeding and, more recently, using the modern tools of genetic engineering and biotechnology. However, the heterogenicity, complex poly-aneuploid genome and susceptibility of sugarcane to different biotic and abiotic stresses represent impediments that require us to pay greater attention to the improvement of the sugarcane crop. Compared to traditional breeding, recent advances in breeding technologies (molecular marker-assisted breeding, sugarcane transformation, genome-editing and multiple omics technologies) can potentially improve sugarcane, especially against environmental stressors. This article will focus on efficient modern breeding technologies, which provide crucial clues for the engineering of sugarcane cultivars resistant to environmental stresses.

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Javed Ashraf, M.R. Foolad (2007)

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High-fidelity CRISPR-Cas9 variants with undetectable genome-wide off-targets

Benjamin P. Kleinstiver, Vikram Pattanayak, Michelle S. Prew … (2016)

CRISPR-Cas9 nucleases are widely used for genome editing but can induce unwanted off-target mutations. Existing strategies for reducing genome-wide off-targets of the broadly used Streptococcus pyogenes Cas9 (SpCas9) are imperfect, possessing only partial or unproven efficacies and other limitations that constrain their use. Here we describe SpCas9-HF1, a high-fidelity variant harboring alterations designed to reduce non-specific DNA contacts. SpCas9-HF1 retains on-target activities comparable to wild-type SpCas9 with >85% of single-guide RNAs (sgRNAs) tested in human cells. Strikingly, with sgRNAs targeted to standard non-repetitive sequences, SpCas9-HF1 rendered all or nearly all off-target events undetectable by genome-wide break capture and targeted sequencing methods. Even for atypical, repetitive target sites, the vast majority of off-targets induced by SpCas9-HF1 were not detected. With its exceptional precision, SpCas9-HF1 provides an alternative to wild-type SpCas9 for research and therapeutic applications. More broadly, our results suggest a general strategy for optimizing genome-wide specificities of other RNA-guided nucleases.