High efficacy full allelic CRISPR/Cas9 gene editing in tetraploid potato

Johansen, Ida Elisabeth; Liu, Ying; Jørgensen, Bodil; Bennett, Eric; Andréasson, Erik; Nielsen, Kåre Lehmann; Blennow, Andreas; Petersen, Bent Larsen

doi:10.1038/s41598-019-54126-w

Cited by 87 publications

(75 citation statements)

References 37 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…CRISPR/Cas9 has become a cutting-edge technology to provide an opportunity for more effective crop improvement than other commonly used methods ( Cong et al, 2013 ; Mali et al, 2013 ; Xie et al, 2015 ; Wang et al, 2016 ; Park et al, 2017 ; Zhang et al, 2018 ; Erpen-Dalla et al, 2019 ; Jiang et al, 2019 ; Johansen et al, 2019 ; Tofazzal, 2019 ; Oh et al, 2020 ; Niu et al, 2020 ; Zhang et al, 2020 ). Regardless of substantial achievements in several crops, the efficiency of the CRISPR/Cas9 system in alfalfa is still challenging ( Meng et al, 2017 ; Gao et al, 2018 ), which necessitate the development of a robust and effective CRISPR/Cas9 genome editing system.…”

Section: Discussionmentioning

confidence: 99%

“…Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) has become the most preferred genome editing tool, enabling continuous success in genetic manipulation of many crops ( Voytas and Gao, 2014 ; Ma et al, 2016 ; Wang et al, 2016 ; Meng et al, 2017 ; Barry et al, 2018 ; Zhang et al, 2018 ; Erpen-Dalla et al, 2019 ; Jiang et al, 2019 ; Johansen et al, 2019 ; Tofazzal, 2019 ; Oh et al, 2020 ; Niu et al, 2020 ; Zhang et al, 2020 ). To date, several polyploid crops have been successfully edited by the gRNA-CRISPR/Cas9 genome editing system, including wheat ( Triticum aestivum ), cotton ( Gossypium hirsutum ), rapeseed ( Brassica napus ), and potato ( Solanum tuberosum ) ( Wang et al, 2014 ; Sun et al, 2015 ; Wang et al, 2016 ; Andersson et al, 2017 ; Cai et al, 2017 ; Zhang et al, 2017 ; Kanazashi et al, 2018 ; Kusano et al, 2018 ; Johansen et al, 2019 ). However, in spite of such substantial genome editing achievements, the use of the CRISPR/Cas9 system in alfalfa is still very challenging compared to the improvements made in other dicot and monocot plants.…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Development of a Highly Efficient Multiplex Genome Editing System in Outcrossing Tetraploid Alfalfa (Medicago sativa)

et al. 2020

View full text Add to dashboard Cite

Alfalfa ( Medicago sativa ) is an outcrossing tetraploid legume species widely cultivated in the world. The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (CRISPR/Cas9) system has been successfully used for genome editing in many plant species. However, the use of CRISPR/Cas9 for gene knockout in alfalfa is still very challenging. Our initial single gRNA-CRISPR/Cas9 system had very low mutagenesis efficiency in alfalfa with no mutant phenotype. In order to develop an optimized genome editing system in alfalfa, we constructed multiplex gRNA-CRISPR/Cas9 vectors by a polycistronic tRNA-gRNA approach targeting the Medicago sativa stay-green ( MsSGR ) gene. The replacement of CaMV35S promoter by the Arabidopsis ubiquitin promoter (AtUBQ10) to drive Cas9 expression in the multiplex gRNA system led to a significant improvement in genome editing efficiency, whereas modification of the gRNA scaffold resulted in lower editing efficiency. The most effective multiplex system exhibited 75% genotypic mutagenesis efficiency, which is 30-fold more efficient than the single gRNA vector. Importantly, phenotypic change was easily observed in the mutants, and the phenotypic mutation efficiency reached 68%. This highly efficient multiplex gRNA-CRISPR/Cas9 genome editing system allowed the generation of homozygous mutants with a complete knockout of the four allelic copies in the T0 generation. This optimized system offers an effective way of testing gene functions and overcomes a major barrier in the utilization of genome editing for alfalfa improvement.

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Development of a Highly Efficient Multiplex Genome Editing System in Outcrossing Tetraploid Alfalfa (Medicago sativa)

et al. 2020

View full text Add to dashboard Cite

show abstract

“…Therefore, selection of the guide RNA sequence will need to be preceded by resequencing of the target gene in the individual genotype to be transformed. In potato, endogenous promoters have been used to greatly increase the efficiency of CRISPR-mediated genome editing ( Liang et al , 2018 ; Johansen et al , 2019 ); similar strategies should be explored in quinoa. Moreover, engineering of wheat with the CRISPR/Cas9 system required codon optimization of the Cas9 sequence and the use of a maize promoter for expression ( Zhang et al , 2019 ).…”

Section: Introductionmentioning

confidence: 99%

Prospects for the accelerated improvement of the resilient crop quinoa

López‐Marqués

Nørrevang

Ache

et al. 2020

Journal of Experimental Botany

View full text Add to dashboard Cite

Abstract Crops tolerant to drought and salt stress may be developed by two approaches. First, major crops may be improved by introducing genes from tolerant plants. For example, many major crops have wild relatives that are more tolerant to drought and high salinity than the cultivated crops, and, once deciphered, the underlying resilience mechanisms could be genetically manipulated to produce crops with improved tolerance. Secondly, some minor (orphan) crops cultivated in marginal areas are already drought and salt tolerant. Improving the agronomic performance of these crops may be an effective way to increase crop and food diversity, and an alternative to engineering tolerance in major crops. Quinoa (Chenopodium quinoa Willd.), a nutritious minor crop that tolerates drought and salinity better than most other crops, is an ideal candidate for both of these approaches. Although quinoa has yet to reach its potential as a fully domesticated crop, breeding efforts to improve the plant have been limited. Molecular and genetic techniques combined with traditional breeding are likely to change this picture. Here we analyse protein-coding sequences in the quinoa genome that are orthologous to domestication genes in established crops. Mutating only a limited number of such genes by targeted mutagenesis appears to be a promising route for accelerating the improvement of quinoa and generating a nutritious high-yielding crop that can meet the future demand for food production in a changing climate.

show abstract

“…Precise genome editing (PGE) has been introduced for starch bioengineering targeting specific genes by mutagenesis, generating non-GM crops. Recently the CRISPR/ Cas9 (clustered regularly interspaced short palindromic repeats/ Cas9) system was used to knock out the granule-bound starch synthase (GBSS) to generate amylose-free potato tuber starch (Clasen et al, 2015;Johansen et al, 2019;Veillet et al, 2019) and combinations of SBEI and SBEII to achieve high-amylose potato starch (Tuncel et al, 2019).…”

Section: Biotechnologies For Starch Bioengineeringmentioning

confidence: 99%

Non‐GMO potato lines, synthesizing increased amylose and resistant starch, are mainly deficient in isoamylase debranching enzyme

Blennow

Skryhan

Tanackovic

et al. 2020

Plant Biotechnology Journal

Self Cite

View full text Add to dashboard Cite

Summary Solanum tuberosum potato lines with high amylose content were generated by crossing with the wild potato species Solanum sandemanii followed by repeated backcrossing to Solanum tuberosum lines. The trait, termed increased amylose (IAm), was recessive and present after three generations of backcrossing into S. tuberosum lines (6.25% S. sandemanii genes). The tubers of these lines were small, elongated and irregular with small and misshaped starch granules and high sugar content. Additional backcrossing resulted in less irregular tuber morphology, increased starch content (4.3%–9.5%) and increased amylose content (29%–37.9%) but indifferent sugar content. The amylose in the IAm starch granules was mainly located in peripheral spots, and large cavities were found in the granules. Starch pasting was suppressed, and the digestion‐resistant starch (RS) content was increased. Comprehensive microarray polymer profiling (CoMPP) analysis revealed specific alterations of major pectic and glycoprotein cell wall components. This complex phenotype led us to search for candidate IAm genes exploiting its recessive trait. Hence, we sequenced genomic DNA of a pool of IAm lines, identified SNPs genome wide against the draft genome sequence of potato and searched for regions of decreased heterozygosity. Three regions, located on chromosomes 3, 7 and 10, respectively, displayed markedly less heterozygosity than average. The only credible starch metabolism‐related gene found in these regions encoded the isoamylase‐type debranching enzyme Stisa1. Decreased expression of mRNA (>500 fold) and reduced enzyme activity (virtually absent from IAm lines) supported Stisa1 as a candidate gene for IAm.

show abstract

High efficacy full allelic CRISPR/Cas9 gene editing in tetraploid potato

Cited by 87 publications

References 37 publications

Development of a Highly Efficient Multiplex Genome Editing System in Outcrossing Tetraploid Alfalfa (Medicago sativa)

Development of a Highly Efficient Multiplex Genome Editing System in Outcrossing Tetraploid Alfalfa (Medicago sativa)

Prospects for the accelerated improvement of the resilient crop quinoa

Non‐GMO potato lines, synthesizing increased amylose and resistant starch, are mainly deficient in isoamylase debranching enzyme

Contact Info

Product

Resources

About