Sorghum (Sorghum bicolor L. Moench) is one of the world's most cultivated cereal crops. Biotechnology approaches have great potential to complement traditional crop improvement. Earlier studies in rice and maize revealed that LIGULELESS-1 (LG1) is responsible for formation of the ligule and auricle, which determine the leaf inclination angle. However, generation and analysis of lg1 mutants in sorghum has so far not been described. Here, we describe CRISPR/Cas9 mediated targeted mutagenesis of LG1 in sorghum and phenotypic changes in mono- and bi-allelic lg1 mutants. Genome editing reagents were co-delivered to sorghum (var. Tx430) with the nptII selectable marker via particle bombardment of immature embryos followed by regeneration of transgenic plants. Sanger sequencing confirmed a single nucleotide insertion in the sgRNA LG1 target site. Monoallelic edited plantlets displayed more upright leaves in tissue culture and after transfer to soil when compared to wild type. T1 progeny plants with biallelic lg1 mutation lacked ligules entirely and displayed a more severe reduction in leaf inclination angle than monoallelic mutants. Transgene-free lg1 mutants devoid of the genome editing vector were also recovered in the segregating T1 generation. Targeted mutagenesis of LG1 provides a rapidly scorable phenotype in tissue culture and will facilitate optimization of genome editing protocols. Altering leaf inclination angle also has the potential to elevate yield in high-density plantings.

CRISPR/Cas9 mediated targeted mutagenesis of LIGULELESS-1 in sorghum provides a rapidly scorable phenotype by altering leaf inclination angle.

CRISPR/Cas9 technology as an innovative approach to enhancing the phytoremediation: Concepts and implications.

Sorghum is a versatile crop with great potential as a sustainable food, feed, and bioenergy source. To mitigate the severely negative impact of climate change and population growth on food and energy security, further elevation of the crops stress tolerance is urgently needed. Genome editing technologies such as CRISPR/Cas have great potential to accelerate functional genomics and crop improvement by supporting targeted modification of almost any crop gene sequence. We describe the recent progress in genome editing of sorghum. In addition, we review remaining challenges and prospects of emerging gene editing technologies for rapid precision breeding of this crop.

CRISPR/Cas-mediated genome editing in sorghum — recent progress, challenges and prospects

Progresses of CRISPR/Cas9 genome editing in forage crops.

A novel in vitro propagation system for West Indian elm [Guazuma ulmifolia Lam. (Malvaceae)]: a valuable medicinal woody species

Napier grass (Cenchrus purpureus Schum.) is one of the highest yielding tropical C4 forage grasses and a promising candidate feedstock for the production of biofuel. Genetic engineering and genome editing have tremendous potential for crop improvement and require the establishment of a transformation protocol. In this study, transgenic Napier grass was generated by introducing a minimal cassette (MC) encoding the antibiotic resistance gene nptII with biolistic gene transfer into embryogenic callus. Paromomycin was superior to geneticin as a selective agent in the selection of transgenic calli. Two high-biomass producing Napier grass accessions were successfully transformed. Southern blot analysis revealed simple transgene integration patterns with 33% of the regenerated transgenic lines having a single insert of the minimal transgene expression cassette. NPTII-ELISA confirmed transgene expression in all transgenic lines. The results of this study will enable the acceleration of Napier grass improvement by genetic transformation or genome editing.

Aalap Parikh

Papers

CRISPR/Cas9 mediated targeted mutagenesis of LIGULELESS-1 in sorghum provides a rapidly scorable phenotype by altering leaf inclination angle.

CRISPR/Cas-mediated genome editing in sorghum — recent progress, challenges and prospects

Generation of Transgenic Napier Grass (Cenchrus purpureus Schum.) Plants by Biolistic Gene Transfer of a Minimal Expression Cassette