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Journal Article Ppublish 25 citations

PubMed: 36964962 DOI: 10.1111/pbi.14042

Multiplex CRISPR/Cas9-mediated mutagenesis of alfalfa FLOWERING LOCUS Ta1 (MsFTa1) leads to delayed flowering time with improved forage biomass yield and quality.

Plant Biotechnol J 21(7):1383-1392

Authors: Tezera W Wolabu, Kashif Mahmood, Ivone Torres Jerez, Lili Cong, Jianfei Yun, Michael Udvardi, Million Tadege, Zengyu Wang, Jiangqi Wen

Affiliations

  • Institute for Agricultural Biosciences, Oklahoma State University, Oklahoma, Ardmore, USA.
  • College of Grassland Science, Qingdao Agricultural University, Qingdao, Shandong, China.
  • Queensland Alliance for Agriculture and Food Innovation, The University of Queensland, St. Lucia, Queensland, Australia.

Abstract

Alfalfa (Medicago sativa L.) is a perennial flowering plant in the legume family that is widely cultivated as a forage crop for its high yield, forage quality and related agricultural and economic benefits. Alfalfa is a photoperiod sensitive long-day (LD) plant that can accomplish its vegetative and reproductive phases in a short period of time. However, rapid flowering can compromise forage biomass yield and quality. Here, we attempted to delay flowering in alfalfa using multiplex CRISPR/Cas9-mediated mutagenesis of FLOWERING LOCUS Ta1 (MsFTa1), a key floral integrator and activator gene. Four guide RNAs (gRNAs) were designed and clustered in a polycistronic tRNA-gRNA system and introduced into alfalfa by Agrobacterium-mediated transformation. Ninety-six putative mutant lines were identified by gene sequencing and characterized for delayed flowering time and related desirable agronomic traits. Phenotype assessment of flowering time under LD conditions identified 22 independent mutant lines with delayed flowering compared to the control. Six independent Msfta1 lines containing mutations in all four copies of MsFTa1 accumulated significantly higher forage biomass yield, with increases of up to 78% in fresh weight and 76% in dry weight compared to controls. Depending on the harvesting schemes, many of these lines also had reduced lignin, acid detergent fibre (ADF) and neutral detergent fibre (NDF) content and significantly higher crude protein (CP) and mineral contents compared to control plants, especially in the stems. These CRISPR/Cas9-edited Msfta1 mutants could be introduced in alfalfa breeding programmes to generate elite transgene-free alfalfa cultivars with improved forage biomass yield and quality.

MeSH

Mutagenesis, Biotechnology, Clustered Regularly Interspaced Short Palindromic Repeats, Detergents, Genes, Biomass

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